Alkaloids and flavonoids from African phytochemicals as potential inhibitors of SARS-Cov-2 RNA-dependent RNA polymerase: an in silico perspective

Corona Virus Disease 2019 (COVID-19) is a pandemic caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Exploiting the potentials of phytocompounds is an integral component of the international response to this pandemic. In this study, a virtual screening through molecular docking analysis was used to screen a total of 226 bioactive compounds from African herbs and medicinal plants for direct interactions with SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). From these, 36 phytocompounds with binding affinities higher than the approved reference drugs (remdesivir and sobosivir), were further docked targeting the active sites of SARS-CoV-2, as well as SARS-CoV and HCV RdRp. A hit list of 7 compounds alongside two positive controls (remdesivir and sofosbuvir) and two negative controls (cinnamaldehyde and Thymoquinone) were further docked into the active site of 8 different conformations of SARS-CoV-2 RdRp gotten from molecular dynamics simulation (MDS) system equilibration. The top docked compounds were further subjected to predictive druglikeness and ADME/tox filtering analyses. Drugable alkaloids (10’–hydroxyusambarensine, cryptospirolepine, strychnopentamine) and flavonoids (usararotenoid A, and 12α-epi-millettosin), were reported to exhibit strong affinity binding and interactions with key amino acid residues in the catalytic site, the divalent-cation–binding site, and the NTP entry channel in the active region of the RdRp enzyme as the positive controls. These phytochemicals, in addition to other promising antivirals such as remdesivir and sofosbuvir, may be exploited towards the development of a cocktail of anti-coronavirus treatments in COVID-19. Experimental studies are recommended to validate these study.

Download Full-text

Identification of Residues Critical for the Interferon Antagonist Function of Langat Virus NS5 Reveals a Role for the RNA-Dependent RNA Polymerase Domain

Journal of Virology ◽

10.1128/jvi.02830-06 ◽

2007 ◽

Vol 81 (13) ◽

pp. 6936-6946 ◽

Cited By ~ 53

Author(s):

Gregory S. Park ◽

Keely L. Morris ◽

Roselyn G. Hallett ◽

Marshall E. Bloom ◽

Sonja M. Best

Keyword(s):

Amino Acids ◽

Rna Polymerase ◽

Nonstructural Protein ◽

Encephalitis Virus ◽

Janus Kinase ◽

Amino Acid Residues ◽

Rna Dependent Rna Polymerase ◽

Stat Signaling ◽

Langat Virus ◽

Tick Borne Encephalitis Virus

ABSTRACT All pathogenic flaviviruses examined thus far inhibit host interferon (IFN) responses by suppressing the Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway. Both Langat virus (LGTV; a member of the tick-borne encephalitis virus serogroup) and Japanese encephalitis virus use the nonstructural protein NS5 to suppress JAK-STAT signaling. However, NS5 is also critical to virus replication, contributing methyltransferase and RNA-dependent RNA polymerase (RdRP) activities. The specific amino acid residues of NS5 involved in IFN antagonism are not known. Here, we demonstrate that the LGTV NS5 JAK-STAT inhibitory domain is contained between amino acids 355 and 735 (of 903), a range which lies within the RdRP domain. Furthermore, we identified two noncontiguous stretches of specific amino acids within the RdRP, 374 to 380 and 624 to 647, as critical for inhibition of JAK-STAT signaling. Despite considerable separation on the linear NS5 sequence, these residues localized adjacent to each other when modeled on the West Nile virus RdRP crystal structure. Due to the general conservation of RdRP structures, these results suggest that the specific residues identified act cooperatively to form a unique functional site on the RdRP responsible for JAK-STAT inhibition. This insight into the mechanism underlying flavivirus IFN evasion strategies will facilitate the design of antiviral therapeutics that potentiate the action of IFN during infection.

Download Full-text

Probing remdesivir nucleotide analogue insertion to SARS-CoV-2 RNA dependent RNA polymerase in viral replication

10.1101/2021.07.12.452099 ◽

2021 ◽

Author(s):

Moises Ernesto Romero ◽

Chunhong Long ◽

Daniel La Rocco ◽

Anusha Mysore Keerthi ◽

Dajun Xu ◽

...

Keyword(s):

Free Energy ◽

Rna Polymerase ◽

Active Site ◽

Energy Barrier ◽

Umbrella Sampling ◽

Dynamics Simulation ◽

Free Energy Barrier ◽

Base Stacking ◽

Rna Dependent Rna Polymerase ◽

Nucleotide Analogue

Remdesivir (RDV) prodrug can be metabolized into a triphosphate form nucleotide analogue (RDV-TP) to bind and insert into the active site of viral RNA dependent RNA polymerase (RdRp) to further interfere with the viral genome replication. In this work, we computationally studied how RDV-TP binds and inserts to the SARS-CoV-2 RdRp active site, in comparison with natural nucleotide substrate adenosine triphosphate (ATP). To do that, we first constructed atomic structural models of an initial binding complex (active site open) and a substrate insertion complex (active site closed), based on high-resolution cryo-EM structures determined recently for SARS-CoV-2 RdRp or non-structural protein (nsp) 12, in complex with accessory protein factors nsp7 and nsp8. By conducting all-atom molecular dynamics simulation with umbrella sampling strategies on the nucleotide insertion between the open and closed state RdRp complexes, our studies show that RDV-TP can bind comparatively stabilized to the viral RdRp active site, as it primarily forms base stacking with the template Uracil nucleotide (at +1), which is under freely fluctuations and supports a low free energy barrier of the RDV-TP insertion (~ 1.5 kcal/mol). In comparison, the corresponding natural substrate ATP binds to the RdRp active site in Watson-Crick base pairing with the template nt, and inserts into the active site with a medium low free energy barrier (~ 2.6 kcal/mol), when the fluctuations of the template nt are well quenched. The simulations also show that the initial base stacking of RDV-TP with the template can be particularly stabilized by motif B-N691, S682, and motif F-K500 with the sugar, base, and the template backbone, respectively. Although the RDV-TP insertion can be hindered by motif-F R555/R553 interaction with the triphosphate, the ATP insertion seems to be facilitated by such interactions. The inserted RDV-TP and ATP can be further distinguished by specific sugar interaction with motif B-T687 and motif-A D623, respectively.

Download Full-text

Computationally repurposed drugs and natural products against RNA dependent RNA polymerase as potential COVID-19 therapies

Molecular Biomedicine ◽

10.1186/s43556-021-00050-3 ◽

2021 ◽

Vol 2 (1) ◽

Author(s):

Sakshi Piplani ◽

Puneet Kumar Singh ◽

David A. Winkler ◽

Nikolai Petrovsky

Keyword(s):

Natural Products ◽

Rna Polymerase ◽

Kinase Inhibitor ◽

Binding Energies ◽

Dynamics Simulation ◽

Rna Dependent Rna Polymerase ◽

Starting Point ◽

Potential Activity ◽

Repurposed Drugs ◽

Multiple Drugs

AbstractRepurposing of existing drugs and drug candidates is an ideal approach to identify new potential therapies for SARS-CoV-2 that can be tested without delay in human trials of infected patients. Here we applied a virtual screening approach using Autodock Vina and molecular dynamics simulation in tandem to calculate binding energies for repurposed drugs against the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). We thereby identified 80 promising compounds with potential activity against SARS-Cov2, consisting of a mixture of antiviral drugs, natural products and drugs with diverse modes of action. A substantial proportion of the top 80 compounds identified in this study had been shown by others to have SARS-CoV-2 antiviral effects in vitro or in vivo, thereby validating our approach. Amongst our top hits not previously reported to have SARS-CoV-2 activity, were eribulin, a macrocyclic ketone analogue of the marine compound halichondrin B and an anticancer drug, the AXL receptor tyrosine kinase inhibitor bemcentinib. Our top hits from our RdRp drug screen may not only have utility in treating COVID-19 but may provide a useful starting point for therapeutics against other coronaviruses. Hence, our modelling approach successfully identified multiple drugs with potential activity against SARS-CoV-2 RdRp.

Download Full-text

Virtual Screening of the Indonesian Medicinal Plant and Zinc Databases for Potential Inhibitors of the RNA-Dependent RNA Polymerase (RdRp) of 2019 Novel Coronavirus

Indonesian Journal of Chemistry ◽

10.22146/ijc.56120 ◽

2020 ◽

Vol 20 (6) ◽

pp. 1430

Author(s):

Muhammad Arba ◽

Andry Nur-Hidayat ◽

Ida Usman ◽

Arry Yanuar ◽

Setyanto Tri Wahyudi ◽

...

Keyword(s):

Binding Energy ◽

Rna Polymerase ◽

Antiviral Drug ◽

Dynamics Simulation ◽

Human Beings ◽

Rna Dependent Rna Polymerase ◽

Energy Prediction ◽

Binding Energy Prediction ◽

Solvation Energies ◽

Novel Coronavirus

The novel coronavirus disease 19 (Covid-19) which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been a pandemic across the world, which necessitate the need for the antiviral drug discovery. One of the potential protein targets for coronavirus treatment is RNA-dependent RNA polymerase. It is the key enzyme in the viral replication machinery, and it does not exist in human beings, therefore its targeting has been considered as a strategic approach. Here we describe the identification of potential hits from Indonesian Herbal and ZINC databases. The pharmacophore modeling was employed followed by molecular docking and dynamics simulation for 40 ns. 151 and 14480 hit molecules were retrieved from Indonesian herbal and ZINC databases, respectively. Three hits that were selected based on the structural analysis were stable during 40 ns, while binding energy prediction further implied that ZINC1529045114, ZINC169730811, and 9-Ribosyl-trans-zeatin had tighter binding affinities compared to Remdesivir. The ZINC169730811 had the strongest affinity toward RdRp compared to the other two hits including Remdesivir and its binding was corroborated by electrostatic, van der Waals, and nonpolar contribution for solvation energies. The present study offers three hits showing tighter binding to RdRp based on MM-PBSA binding energy prediction for further experimental verification.

Download Full-text

Analysis of tombusvirus revertants to identify essential amino acid residues within RNA-dependent RNA polymerase motifs

Journal of General Virology ◽

10.1099/vir.0.80625-0 ◽

2005 ◽

Vol 86 (3) ◽

pp. 823-826 ◽

Cited By ~ 6

Author(s):

K. Boonrod ◽

S. Chotewutmontri ◽

D. Galetzka ◽

G. Krczal

Keyword(s):

Rna Polymerase ◽

Nicotiana Benthamiana ◽

Essential Amino Acid ◽

Selection Pressure ◽

Rna Binding ◽

Amino Acid Residues ◽

Rna Dependent Rna Polymerase ◽

Viral Rdrp ◽

In Trans ◽

Residue Position

The RNA-dependent RNA polymerase (RdRp) of Tomato bushy stunt virus (TBSV) contains an arginine- and proline-rich (RPR) motif. This motif functions as an RNA-binding domain and is essential for tombusvirus replication. A mutant carrying three arginine substitutions in this motif rendered the virus unable to replicate in Nicotiana benthamiana plants and protoplasts. When the replicase function was provided in trans, by expressing the TBSV RdRp in N. benthamiana plants, an infectious variant could be isolated. Sequence analysis showed that only the substituted glycine residue (position 216) had reverted to arginine; all other substitutions remained unchanged. This finding suggested that strong selection pressure is active to maintain necessary sequences of the viral RdRp and that the analysis of revertants may help to identify essential viral functions.

Download Full-text

Shogaol, Bisdemethoxycurcumin, and Curcuminoid: Potential Zingiber Compounds Against COVID-19

Biointerface Research in Applied Chemistry ◽

10.33263/briac115.1286912876 ◽

2021 ◽

Vol 11 (5) ◽

pp. 12869-12876

Keyword(s):

Bioactive Compounds ◽

Binding Sites ◽

Active Sites ◽

Zingiber Officinale ◽

Amino Acid Residues ◽

Curcuma Zedoaria ◽

Rna Dependent Rna Polymerase ◽

Protein Amino Acid ◽

Discovery Studio ◽

Global Pandemic

Coronavirus disease (COVID-19) is a global pandemic in the world. Some treatments, including vaccines and potential drugs, are still developed. This study investigated the bioactive compounds of Zingiber officinale, Kaempferia rotunda, and Curcuma zedoaria as a potential inhibitor for ACE2 and RdRP proteins. Molecular docking was used for screening the bioactive compounds as ACE2 and RdRP inhibitors. Shogaol (CID 5281794), zingerone (CID 31211), chalcone (CID 637760), Ar-turmerone (CID 558221), bisdemothxycurcumin (CID 5315472), and curcuminoid (CID 101341353) interacted with angiotensin-converting enzyme receptor-2/ACE2 (PDB ID 2xd3) and RNA dependent RNA polymerase/RdRP (PDB ID 6xqb), then analyzed using Discovery studio v.19 program. Shogaol, zingerone, chalcone, ar-turmerone, bisdemethoxycurcumin, and curcuminoid bound to ACE2 and RdRP protein in some active sites. Zingerone, chalcone, and ar-turmerone are attached to the ACE-2 and then RdRP protein in similar active sites, suggesting those compounds stabilize the complex ACE-2 and RdRP protein. Shogaol interacted with the RdRP and ACE2 protein amino acid residues in the Shogaol-RdRP+ACE2 complex, indicating shogaol blocks the RdRP-ACE2 interaction. Then, bisdemethoxycurcumin and curcuminoid change the binding sites of ACE2 and RdRP protein when both compounds are bound to RdRP protein. This study suggested that shogaol, bisdemethoxycurcumin, and curcuminoid are potential drugs for COVID-19 prevention.

Download Full-text

Potential Phytochemical Inhibitors of the Coronavirus RNA Dependent RNA Polymerase: A Molecular Docking Study

10.21203/rs.3.rs-35334/v1 ◽

2020 ◽

Cited By ~ 1

Author(s):

Ardra. P ◽

Prachi Singh ◽

Hariprasad VR ◽

Babu UV ◽

Mohamed Rafiq ◽

...

Keyword(s):

Molecular Docking ◽

Rna Polymerase ◽

Metal Ion ◽

Docking Study ◽

Nucleoside Triphosphate ◽

Root Extract ◽

Amino Acid Residues ◽

Molecular Docking Study ◽

Rna Dependent Rna Polymerase ◽

Phytochemical Compounds

Abstract The COVID-19 disease that originated in China by the end of 2019 has now become a pandemic and has affected 216 countries as on 08 June 2020. RNA dependent RNA polymerase (RdRp), the core enzyme in the multiprotein replicase-transcriptase complex of coronaviruses, serves as a classical target for inhibiting the coronavirus infectivity. In this study we performed molecular docking of sixty-nine different phytochemical compounds from various herbs with RdRp of both SARS-CoV-2 and its predecessor SARS-CoV. Our results show that various phytochemical constituents from Withania somnifera root extract, Hyssopus officinalis and Camellia sinensis leaf extract have high binding affinity towards RdRps and are comparable to the small molecule drug remdesivir. Their binding interactions reveal that they bind to the amino acid residues involved in nucleoside triphosphate (NTP) entry and recognition site and metal ion cofactor chelating site of both SARS-CoV-2 and SARS-CoV. Hence they are different from the classical nucleotide analog inhibitors of RdRp. This study paves a quick platform for development of targeted therapy using phytochemicals for COVID-19 and other potential SARS coronavirus related outbreaks in future.

Download Full-text

The Binding of Remdesivir to SARS-CoV-2 RNA-Dependent RNA polymerase May Pave The Way Towards the Design of Potential Drugs for COVID-19 Treatment.

Current Pharmaceutical Biotechnology ◽

10.2174/1389201021666201027154833 ◽

2020 ◽

Vol 21 ◽

Author(s):

Clement Agoni ◽

Mahmoud E.S. Soliman

Keyword(s):

Molecular Dynamics ◽

Rna Polymerase ◽

Vaccine Development ◽

Treatment Options ◽

Pharmacophore Model ◽

Dynamics Simulation ◽

Active Site Residues ◽

Rna Dependent Rna Polymerase ◽

Hydrophobic Region ◽

Virus Rna

Aim: We seek to provide an understanding of the binding mechanism of Remdesivir, provide structural and conformational implications on SARS-CoV-2 virus RNA-dependent RNA polymerase upon its binding and identify its crucial pharmacophoric moieties. Background: The coronavirus disease of 2019 (COVID-19) pandemic has infected over a million people, with over 65,000 deaths as of the first quarter of 2020. The current limitation of effective treatment options with no approved vaccine or targeted therapeutics for the treatment of COVID-19 has posed serious global health threats. This has necessitated several drug and vaccine development efforts across the globe. To date, the farthest in the drug development pipeline so far is Remdesivir. Objectives: We perform molecular dynamics simulation, quantify the energy contributions of binding site residues using per-residue energy decomposition calculations, and subsequently generate a pharmacophore model for the identification of potential SARS-CoV-2 virus RNA-dependent RNA polymerase inhibitors. Methods: Integrative molecular dynamics simulations and thermodynamic calculations coupled with advanced postmolecular dynamics analysis techniques were employed. Results: Our analysis showed that the modulatory activity of Remdesivir is characterized by an extensive array of highaffinity and consistent molecular interactions with specific active site residues that anchor Remdemsivir within the binding pocket for efficient binding. These residues are ASP452, THR456, ARG555, THR556, VAL557, ARG624, THR680, SER681, and SER682. Results also showed that Remdesivir binding, induces minimal individual amino acid perturbations, subtly interferes with deviations of C-α atoms and restricts the systematic transition of SARS-CoV-2 RNA-dependent RNA polymerase from the “buried” hydrophobic region to the “surface-exposed” hydrophilic region. We also mapped a pharmacophore model based on observed high-affinity interactions with SARS-CoV-2 virus RNA-dependent RNA polymerase, which showcased the crucial functional moieties of Remdesivir and was subsequently employed for virtual screening. Conclusion: The structural insights and the optimized pharmacophoric model provided would augment the design of improved analogs of Remdesivir that could expand treatment options for COVID-19.

Download Full-text

Mechanism of Inhibition of Ebola Virus RNA-Dependent RNA Polymerase by Remdesivir

Viruses ◽

10.3390/v11040326 ◽

2019 ◽

Vol 11 (4) ◽

pp. 326 ◽

Cited By ~ 179

Author(s):

Egor Tchesnokov ◽

Joy Feng ◽

Danielle Porter ◽

Matthias Götte

Keyword(s):

Rna Polymerase ◽

Rna Synthesis ◽

Ebola Virus ◽

Virus Disease ◽

Chain Termination ◽

Mitochondrial Rna ◽

Long Distance ◽

Rna Dependent Rna Polymerase ◽

Nucleotide Analogue ◽

Nucleotide Incorporation

Remdesivir (GS-5734) is a 1′-cyano-substituted adenosine nucleotide analogue prodrug that shows broad-spectrum antiviral activity against several RNA viruses. This compound is currently under clinical development for the treatment of Ebola virus disease (EVD). While antiviral effects have been demonstrated in cell culture and in non-human primates, the mechanism of action of Ebola virus (EBOV) inhibition for remdesivir remains to be fully elucidated. The EBOV RNA-dependent RNA polymerase (RdRp) complex was recently expressed and purified, enabling biochemical studies with the relevant triphosphate (TP) form of remdesivir and its presumptive target. In this study, we confirmed that remdesivir-TP is able to compete for incorporation with adenosine triphosphate (ATP). Enzyme kinetics revealed that EBOV RdRp and respiratory syncytial virus (RSV) RdRp incorporate ATP and remdesivir-TP with similar efficiencies. The selectivity of ATP against remdesivir-TP is ~4 for EBOV RdRp and ~3 for RSV RdRp. In contrast, purified human mitochondrial RNA polymerase (h-mtRNAP) effectively discriminates against remdesivir-TP with a selectivity value of ~500-fold. For EBOV RdRp, the incorporated inhibitor at position i does not affect the ensuing nucleotide incorporation event at position i+1. For RSV RdRp, we measured a ~6-fold inhibition at position i+1 although RNA synthesis was not terminated. Chain termination was in both cases delayed and was seen predominantly at position i+5. This pattern is specific to remdesivir-TP and its 1′-cyano modification. Compounds with modifications at the 2′-position show different patterns of inhibition. While 2′-C-methyl-ATP is not incorporated, ara-ATP acts as a non-obligate chain terminator and prevents nucleotide incorporation at position i+1. Taken together, our biochemical data indicate that the major contribution to EBOV RNA synthesis inhibition by remdesivir can be ascribed to delayed chain termination. The long distance of five residues between the incorporated nucleotide analogue and its inhibitory effect warrant further investigation.

Download Full-text