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A new proposed mechanism of some known drugs
targeting the SARS-CoV-2 spike glycoprotein using
molecular docking
Tarek Moussa ( tarekmoussa@yahoo.com )
Cairo University
Nevien Sabry
National Research Centre
Research Article
Keywords: SARS-Co-2, COVID-19, molecular docking, oseltamivir, hydroxychloroquine
Posted Date: November 13th, 2020
DOI: https://doi.org/10.21203/rs.3.rs-105677/v1
License: This work is licensed under a Creative Commons Attribution 4.0 International License.
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Abstract
COVID-19 is caused by the novel enveloped beta-coronavirus with a genomic RNA closely related to severe
acute respiratory syndrome-corona virus (SARS-CoV) and is named coronavirus 2 (SARS-CoV-2). The
receptor binding domain (RBD) of the S-protein interacts with the human ACE-2 receptor that enables the
initiation of viral entry. Hence, blocking the S-protein interactions by means of synthetic compounds mark
the pivotal step for targeting SARS-CoV-2. Most of the six compounds were observed to t nicely with
specic noncovalent interactions, including H bonds, electrostatic, Van der Waals and hydrophobic bonds
(pi and sigma bonds). Oseltamivir was found to be the most strongly interacting with the RBD, exhibiting
high values of full tness and low free energy of binding. it formed multiple noncovalent bonds in the
region of the active site. Hydroxychloroquine also demonstrated high binding anity in the solvent
accessbility state and t nicely into the active pocket of the S-protein. The results revealed that these
compounds could be potent inhibitors of S-protein that could, to some extent, block its interaction with
ACE-2. It is obvious from the 3D structure of SARS-CoV-2 spike protein was changed with the interaction of
different drugs, which led to the unsuitability to bind ACE2 receptor. Hence, laboratory studies elucidating
the action of these compounds on SARS-CoV-2 are warranted for clinical assessments. Chloroquine,
hydroxychloroquine and oseltamivir interacted well with the receptor binding domain of S-protein via
noncovalent interactions and recommended as excellent candidates for COVID-19.
Introduction
Coronavirus disease 2019 (COVID-19) occurred sporadically in Wuhan City, China in December 2019, then
quickly spread throughout China and the entire world. Severe acute respiratory syndrome-corona virus 2
(SARS-CoV-2) is one the most infectious evolving current pathogen, causing severe respiratory illness and
morbidity[1–3]. In the absence of drugs or vaccines, the cornerstone for preventing the spread of infection
lies in adopting good personal hygiene, maintaining social distancing, restricting travel and the use of
potentially effective natural products and therapeutics[4,5]. Many articial and natural compounds have
been researched to combat SARS-CoV-2. Most target its spike (S) glycoprotein, which facilitates host cell
entry, primarily by binding to the host angiotensin-converting enzyme 2 (ACE 2) receptor, present on the
surfaces of macrophages, lymphocytes and other immune cells[6–8]. This phenomenon is specic to the
receptor binding domain (RBD) of the S-protein that spans from 326-580 amino acids and has unique
sites for interaction with ACE-2[9–11].
Chloroquine and hydroxychloroquine, which have been used for malaria prevention and treatment for
decades, as well as for the treatment and management of chronic inammatory diseases such as
systemic lupus erythematosus and rheumatoid arthritis, have gained signicant focus as possible
therapies[12–14].Lopinavir is an antiretroviral protease inhibitor used in the treatment of HIV-1 infection
combined with other antiretrovirals[15]. Oseltamiviris used in the treatment of theu virus(inuenza),
ameliorating the symptoms (such as cough, stuffy nose,fever/chills, sore throat, tiredness, aches) and
shortens the recovery time within 1-2 days. Darunavir is anantiretroviral medication that isused in treating
and preventingHIV/AIDS. It is generally recommended to be used with other antiretrovirals[16,17].
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Ibuprofenis a nonsteroidal anti-inammatory drug(NSAID) class medication that is used to
treatpain,fever, andinammation, which includespainful periods of menstruation,migraines,
andrheumatoid arthritis[18].
In this research, we assessed the interaction of Chloroquine, hydroxychloroquine, lopinavir, oseltamivir,
darunavir and ibuprofen with the active site of the RBD of the S protein, using Autodock, Swiss dock and
Discovery Studio tools.
Methods
Preparation of receptor protein and active site prediction
Based on the literature, the cryo-electron microscopy structure of SARS-CoV-2 S protein (PDB ID: 6VSB)
was rst analyzed for ligand interacting sites[19,20]. Spike protein is a homotrimer, so we chose chain A
for the analysis containing all the respective domains. The active site of the target protein was predicted
by using binding site module using Discovery studio[21,22]. The functional active site was present in the
receptor binding domain (Leu 335 to Gly 526), which also comprised the specic active site region or loop
region (NAG of RCSB: 6VSB, chain A) (Figure 1). The active site residue ASN343 was chosen as the centre
of the grid with the following coordinates: spacing: 0.753 Å, XYZ values: 76 126 76, center coordinates:
-36.336 23.128 21.195. For site-specic docking, the X-ray diffraction 3D structure of the specic RBD of S
protein (PDBID: 6M0J) with resolution 2.32.Å was chosen for interaction with the ligands. The above
coordinates were assigned to the 3D structure followed by docking analysis. The 3D structure was energy
minimized in solvent medium in Swiss PDB viewer to a free energy level of E= -10990.533 KJ/mol using
inbuilt Gromos96 algorithm to obtain the most stable structure of the protein for docking[23,24].
Ligand preparation
The PDB format of the 3D structure of all the ligands were obtained from the drug bank
https://www.drugbank.ca/drugs and PubChem https://pubchem.ncbi.nlm.nih.gov/. Chloroquine,
hydroxychloroquine, darunavir, lopinavir, oseltamivir, remdesivir and ibuprofen were used as ligands
(Figure 2). The ligands were prepared for docking in the Ligand Preparation tool of the Discovery studio.
The root was detected for each ligand, which was eventually saved in PDBQT les similar to that of the
receptor protein.
Molecular docking and post scoring analysis
The X-ray diffraction structure of the S protein of SARS-CoV-2 (PDBID: 6M0J, 30 Å) was used for the
docking analysis (Figure 1). Site specic docking was performed at the active site of the protein prepared
using DS studio client 3.5 as described earlier. Then, all the water molecules and heteroatoms were
removed in the Autodock tool. Hydrogen atoms were added to the model based on an explicit all atom
model. Kollmann charges were added for ensuing interaction with the ligands, and the model was energy
minimized. Both the receptor spike protein and the ligands were processed in Autodock tools as described
above and converted to PDBQT format. Eight sets of docking poses were exhaustively performed; each set
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of Autodock Vina produced 9 conformations; among them, the best pose with RMSD = 0 and lowest free
energy of binding was chosen for further analysis. The interacting sites were further visualized in
Discovery Studio Visualizer and the 2D interaction and nature and types of bonds were determined.
Finally, rescoring of the docked conformations was performed in Swiss dock program with the same
attributes for the receptor protein and ligands for better clarity and renement of ligand protein
interactions. The full tness value was obtained for the best docked site and analyzed with that of
Autodock results.
Results
Receptor protein structure and analysis
The 3D conformer of the RBD of S-protein was exposed to forceeld for energy minimization in the solvent
state. The energy was found to be -10990.63 Kcal/mol. Secondary structure analysis using PROMOTIF
revealed that the RBD mainly comprised beta strands (25.3%) and few alpha helices (11%), in addition to
beta turns, helix-turn helix, beta sheets, and gamma strands (63.7%). The active site center represented
residue ASN 343 and formed a loop-like structure. Hydrophobicity-hydrophilicity analysis of the RBD
residues using Peptide 2.0 software revealed that neutral and hydrophobic amino acids comprised 44.85%
an 38.14%, respectively of the total peptides
(https://www.peptide2.com/N_peptide_hydrophobicity_hydrophilicity.php), which indicated lower water
solubility and more non polar interactions with different compounds. Further analysis using the complete
homotrimeric S-protein of SARS-CoV-2 also revealed that the total protein comprised of 42.14% and 41.1%
of hydrophobic and neutral amino acids, suggesting that S-protein is mainly hydrophobic and can interact
with non-polar compounds (Figure 3).
Docking analysis
The interaction of chloroquine with the RBD was moderately strong (ΔG = -5.5 Kcal/mol) with the specic
interacting residues positioned near the active sites (TRP 436, LEU 441). Noncovalent interactions
comprised 2 hydrogen bonds, and 5 hydrophobic bonds between the ligand and the receptor residues. The
ligand t into the binding pocket of the receptor with a full tness of -770.49 (Table 1, Figure 4).
The interaction of hydroxychloroquine was strong (ΔG = -5.8 Kcal/mol) relative to other compounds, with
several interacting residues near the active site region, forming 4 hydrogen bonds and 5 hydrophobic
interactions. This ligand t into the central binding pocket of the RBD with a full tness of -797.05 and
conferred maximum stability (Table 2, Figure 5).
Oseltamivir ligand interacted well with the RBD with a binding energy of ΔG = -5.7 Kcal/mol and full
tness of -794.93. Several interacting residues were in the active site (LEU 517, PHE 464 and THR 430)
and formed two hydrogen bonds and four hydrophobic bonds (Table 3, Figure 6).
The ligand darunavir interacted less with the RBD with only 1 reside (THR430) forming 2 hydrogen bonds
(Table 4, Figure 7) with tness of -783.37 and binding energy of ΔG = -5.5 Kcal/mol.
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The ligand lopinavir did not interact with the RBD residues using either docking approach; only 1 residue
(SER 469) formed hydrogen bond near the active site (Table 5, Figure 8) with low tness of -677.139 and
ΔG = -3.6 Kcal/mol.
Ibuprofen also interacted less with the receptor protein. Although it had low binding energy compared to
other ligands (ΔG = -6.1 Kcal/mol), the interacting residues were less, and all formed weaker hydrophobic
bonds that signied low interaction with the receptor (Table 6, Figure 9).
Discussion
Several medical and molecular trials involving many natural compounds and synthetic compounds used
for medication of other diseases and molecular trials are being pursued[25]. Repurposing previously used
drugs with health-promoting effects confer several advantages, including reduced costs, faster regulatory
approvals and immediate eld trials. Until a vaccine is developed, some treatment is imperative to reduce
COVID19 mortality and morbidity.
Multiple studies indicate that repurposing chloroquine and hydroxychloroquine can effectively inhibit
Coronaviridae infection (including SARS and SARS-CoV-2)
in vitro
[26–29]. Preliminary conicting clinical
evidence from studies in China and France have put the brakes on their ongoing clinical trials[30,31]. Our
ndings suggested that the interaction of chloroquine and hydroxychloroquine was moderately strong
with residues of the RBD of S-protein and tting well into the binding pocket of the receptor. Therefore,
these compounds could also be potent inhibitors of the S-protein function and should be further evaluated
in clinical trials. Our study indicates that oseltamivir could be useful for combating COVID-19, in contrast
to the study from Wuhan, which observed no positive results[32]. Several clinical trials are testing the
ecacy or oseltamivir in treating COVID-19. Oseltamivir is also used in many combinations in clinical
trials, such as with chloroquine and avipiravir[33]. Our ndings were agreement with darunavir’s antiviral
activity
in vitro
against a clinical isolate from a SARS-CoV-2 patient, where darunavir showed no activity
against SARS-CoV-2 and the data did not support the darunavir use for COVID-19 treatment[34]. Our
nding regarding Lopinavir was in agreement with the study on hospitalized adult patients with severe
SARS-CoV-2, where no benet was observed beyond standard treatment with lopinavir–
ritonavir[35].Similarly, our results for ibuprofen indicated that it may make symptoms worse in COVID-19
patients
38
. They conrmed that the binding of coronaviruses with angiotensin-converting enzyme-2, and
the bioavailability of angiotensin-converting enzyme-2 will be increased by the administration of
ibuprofen, thus enhancing and potentiating the process of coronavirus infection[36].
Overall, the drugs used in this study such as hydroxychloroquine, chloroquine and oseltamivir conferred
highest interactions with the RBD of S-protein of SARS-CoV-2and they are promising as anti-SARS-CoV-2
compounds. Further laboratory studies and eld-based clinical trials are warranted to further extrapolate
the antiviral outcomes of these compounds.
Conclusions
