Hydrogen Bonding (Base Pairing) in Antiviral Activity.

Base pairing based on hydrogen bonding has, since its inception, been crucial in the antiviral activity of arabinosyladenine, 2'-deoxyuridines (i.e., IDU, TFT, BVDU), acyclic nucleoside analogues (i.e., acyclovir) and nucleoside reverse transcriptase inhibitors (NRTIs). Base pairing based on hydrogen bonding also plays a key role in the mechanism of action of various acyclic nucleoside phosphonates (ANPs) such as adefovir, tenofovir, cidofovir and O-DAPYs, thus explaining their activity against a wide array of DNA viruses (human hepatitis B virus (HBV), human immunodeficiency (HIV) and human herpes viruses (i.e., human cytomegalovirus)). Hydrogen bonding (base pairing) also seems to be involved in the inhibitory activity of Cf1743 (and its prodrug FV-100) against varicella-zoster virus (VZV) and in the activity of sofosbuvir against hepatitis C virus and that of remdesivir against SARS-CoV-2 (COVID-19). Hydrogen bonding (base pairing) may also explain the broad-spectrum antiviral effects of ribavirin and favipiravir. This may lead to lethal mutagenesis (error catastrophe), as has been demonstrated with molnutegravir in its activity against SARS-CoV-2.

Elucidating the Role of Noncovalent Interactions in Favipiravir, a Drug Active against Various Human RNA Viruses; a 1H-14N NQDR/Periodic DFT/QTAIM/RDS/3D Hirshfeld Surfaces Combined Study.

Favipiravir (6-fluoro-3-hydroxypyrazine-2-carboxamide, FPV), an active pharmaceutical component of the drug discovered and registered in March 2014 in Japan under the name Avigan, with an indication for pandemic influenza, has been studied. The study of this compound was prompted by the idea that effective processes of recognition and binding of FPV to the nucleic acid are affected predominantly by the propensity to form intra- and intermolecular interactions. Three nuclear quadrupole resonance experimental techniques, namely 1H-14N cross-relaxation, multiple frequency sweeps, and two-frequency irradiation, followed by solid-state computational modelling (density functional theory supplemented by the quantum theory of atoms in molecules, 3D Hirshfeld Surfaces, and reduced density gradient) approaches were applied. The complete NQR spectrum consisting of nine lines indicating the presence of three chemically inequivalent nitrogen sites in the FPV molecule was detected, and the assignment of lines to particular sites was performed. The description of the nearest vicinity of all three nitrogen atoms was used to characterize the nature of the intermolecular interactions from the perspective of the local single atoms and to draw some conclusions on the nature of the interactions required for effective recognition and binding. The propensity to form the electrostatic N-H···O, N-H···N, and C-H···O intermolecular hydrogen bonds competitive with two intramolecular hydrogen bonds, strong O-H···O and very weak N-H···N, closing the 5-member ring and stiffening the structure, as well as π···π and F···F dispersive interactions, were analysed in detail. The hypothesis regarding the similarity of the interaction pattern in the solid and the RNA template was verified. It was discovered that the -NH2 group in the crystal participates in intermolecular hydrogen bonds N-H···N and N-H···O, in the precatalytic state only in N-H···O, while in the active state in N-H···N and N-H···O hydrogen bonds, which is of importance to link FVP to the RNA template. Our study elucidates the binding modes of FVP (in crystal, precatalytic, and active forms) in detail and should guide the design of more potent analogues targeting SARS-CoV-2. Strong direct binding of FVP-RTP to both the active site and cofactor discovered by us suggests a possible alternative, allosteric mechanism of FVP action, which may explain the scattering of the results of clinical trials or the synergistic effect observed in combined treatment against SARS-CoV-2.

The evolution of the spike protein and hACE2 interface of SARS-CoV-2 omicron variants determined by hydrogen bond formation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first detected in December 2019. As of mid-2021, the delta variant was the primary type; however, in January 2022, the omicron (BA.1) variant rapidly spread and became the dominant type in the United States. In June 2022, its subvariants surpassed previous variants in different temporal and spatial situations. To investigate the high transmissibility of omicron variants, we assessed the complex of spike protein 1 receptor-binding domain (S1RBD) and human angiotensin-converting enzyme 2 (hACE2) from the Protein Data Bank (6m0j, 7a91, 7mjn, 7v80, 7v84, 7v8b, 7wbl and 7xo9) and directly mutated specific amino acids to simulate several variants, including variants of concern (alpha, beta, gamma, delta), variants of interest (delta plus, epsilon, lambda, mu, mu without R346K) and omicron variants (BA.1, BA.2, BA.2.12.1, BA.4, BA.5). Molecular dynamics (MD) simulations for 100 ns under physiological conditions were then performed. We found that the omicron S1RBD-hACE2 complexes become more compact with increases in hydrogen-bond interactions at the interface, which is related to the transmissibility of SARS-CoV-2. Moreover, the relaxation time of hydrogen bonds is relatively short among the omicron variants, which implies that the interface conformation alterations are fast. From the molecular perspective, PHE486 and TYR501 in omicron S1RBDs need to involve hydrogen bonds and hydrophobic interactions on the interface. Our study provides structural features of the dominant variants that explain the evolution trend and their increased contagiousness and could thus also shed light on future variant changes.

Hydrogen bonding penalty used for virtual screening to discover potent inhibitors for Papain-Like cysteine proteases of SARS-CoV-2.

The Papain-Like proteases (PLpro) of SARS-CoV-2 play a crucial role in viral replication and the formation of nonstructural proteins. To find available inhibitors, the 3D structure of PLpro of SARS2 was obtained by homologous modelling, and we used this structure as a target to search for inhibitors through molecular docking and MM/GBSA binding free energy rescoring. A novel hydrogen bonding penalty was applied to the screening process, which meanwhile took desolvation into account. Finally, 61 compounds were acquired and 4 of them with IC50 at micromolar level tested in vitro enzyme activity assay, which includes clinical drugs tegaserod. Considering the importance of crystal water molecules, the 4 compounds were re-docked and considered bound waters in the active site as a part of PLpro. The binding modes of these 4 compounds were further explored with metadynamics simulations. The hits will provide a starting point for future key interactions identified and lead optimization targetting PLpro.

Graphs of Hydrogen-Bond Networks to Dissect Protein Conformational Dynamics.

Dynamic hydrogen bonds and hydrogen-bond networks are ubiquitous in proteins and protein complexes. Functional roles that have been assigned to hydrogen-bond networks include structural plasticity for protein function, allosteric conformational coupling, long-distance proton transfers, and transient storage of protons. Advances in structural biology provide invaluable insights into architectures of large proteins and protein complexes of direct interest to human physiology and disease, including G Protein Coupled Receptors (GPCRs) and the SARS-Covid-19 spike protein S, and give rise to the challenge of how to identify those interactions that are more likely to govern protein dynamics. This Perspective discusses applications of graph-based algorithms to dissect dynamical hydrogen-bond networks of protein complexes, with illustrations for GPCRs and spike protein S. H-bond graphs provide an overview of sites in GPCR structures where hydrogen-bond dynamics would be required to assemble longer-distance networks between functionally important motifs. In the case of spike protein S, graphs identify regions of the protein where hydrogen bonds rearrange during the reaction cycle and where local hydrogen-bond networks likely change in a virus variant of concern.

A Competition between Hydrogen, Stacking, and Halogen Bonding in N-(4-((3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)selanyl)phenyl)acetamide: Structure, Hirshfeld Surface Analysis, 3D Energy Framework Approach, and DFT Calculation.

N-(4-((3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)selanyl)phenyl)acetamide (5), C19H15NO3Se, was prepared in two steps from 4,4'-diselanediyldianiline (3) via reduction and subsequent nucleophilic reaction with 2-methyl-3-bromo-1,4-naphthalenedione, followed by acetylation with acetic anhydride. The cytotoxicity was estimated against 158N and 158JP oligodendrocytes and the redox profile was also evaluated using different in vitro assays. The technique of single-crystal X-ray diffraction is used to confirm the structure of compound 5. The enantiopure 5 crystallizes in space group P21 with Flack parameter 0.017 (8), exhibiting a chiral layered absolute structure. Molecular structural studies showed that the crystal structure is foremost stabilized by N-H···O and relatively weak C-H···O contacts between molecules, and additionally stabilized by weak C-H···π and Se···N interactions. Hirshfeld surface analysis is used to quantitatively investigate the noncovalent interactions that stabilize crystal packing. Framework energy diagrams were used to graphically represent the stabilizing interaction energies for crystal packing. The analysis of the energy framework shows that the interactions energies of and C-H···π and C-O···π are primarily dispersive and are the crystal's main important forces. Density functional theory (DFT) calculations were used to determine the compound's stability, chemical reactivity, and other parameters by determining the HOMO-LUMO energy differences. The determination of its optimized surface of the molecular electrostatic potential (MEP) was also carried out. This study was conducted to demonstrate both the electron-rich and electron-poor sites.

Synthesis, structural analysis, and docking studies with SARS-CoV-2 of a trinuclear zinc complex with N-phenylanthranilic acid ligands.

The structure of a trinuclear zinc complex, hexakis(µ2-2-anilinobenzoato)diaquatrizinc(II), [Zn2(C13H10NO2)6(H2O)2] or (NPA)6Zn3(H2O)2 (NPA is 2-anilinobenzoate or N-phenylanthranilate), is reported. The complex crystallizes in the triclinic space group P-1 and the central ZnII atom is located on an inversion center. The NPA ligand is found to coordinate via the carboxylate O atoms with unique C-O bond lengths that support an unequal distribution of resonance over the carboxylate fragment. The axial H2O ligands form hydrogen bonds with neighboring molecules that stabilize the supramolecular system in rigid straight chains, with an angle of 180° along the c axis. π stacking is the primary stabilization along the a and b axes, resulting in a highly ordered supramolecular structure. Docking studies show that this unique supramolecular structure of a trinuclear zinc complex has potential for binding to the main protease (Mpro) in SARS-CoV-2 in a different location from Remdesivir, but with a similar binding strength.

Predicting spike protein NTD mutations of SARS-CoV-2 causing immune evasion by molecular dynamics simulations.

The coronavirus disease 2019 (COVID-19) pandemic was caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Among all the potential targets studied for developing drugs and antibodies, the spike (S) protein is the most striking one, which is on the surface of the virus. In contrast with the intensively investigated immunodominant receptor-binding domain (RBD) of the protein, little is known about the neutralizing antibody binding mechanisms of the N-terminal domain (NTD), let alone the effects of NTD mutations on antibody binding and thereby the risk of immune evasion. Based on 400 ns molecular dynamics simulation for 11 NTD-antibody complexes together with other computational approaches in this study, we investigated critical residues for NTD-antibody binding and their detailed mechanisms. The results show that 36 residues on the NTD including R246, Y144, K147, Y248, L249 and P251 are critically involved in the direct interaction of the NTD with many monoclonal antibodies (mAbs), indicating that the viruses harboring these residue mutations may have a high risk of immune evasion. Binding free energy calculations and an interaction mechanism study reveal that R246I, which is present in the Beta (B.1.351/501Y.V2) variant, may have various impacts on current NTD antibodies through abolishing the hydrogen bonds and electrostatic interaction with the antibodies or affecting other interface residues. Therefore, special attention should be paid to the mutations of these key residues in future antibody and vaccine design and development.

Aromatic Cadinane Sesquiterpenoids from the Fruiting Bodies of Phellinus pini Block SARS-CoV-2 Spike-ACE2 Interaction.

The ongoing COVID-19 global pandemic caused by SARS-CoV-2 inspires the development of effective inhibitors to block the SARS-CoV-2 spike-ACE2 interaction. A chemical investigation on the fruiting bodies of Phellinus pini led to the isolation of five aromatic cadinane sesquiterpenoids including four new ones, named piniterpenoids A-D (1-4), as well as three known lignans. Their structures were determined by extensive spectroscopic analysis including HRMS and 1D and 2D NMR. All of the aromatic cadinane sesquiterpenoids inhibited the SARS-CoV-2 spike-ACE2 interaction, with IC50 values ranging from 64.5 to 99.1 µM. A molecular docking study showed the disruption of the interaction of compound 1 via hydrogen interactions with Arg403, Asp405, and Arg408 of SARS-CoV-2 RBD and Arg393 and His34 residues of ACE2. These results suggested that aromatic cadinane sesquiterpenoids might be useful in developing agents for COVID-19.

SARS-CoV-2 Omicron variant: Antibody evasion and cryo-EM structure of spike protein-ACE2 complex.

The newly reported Omicron variant is poised to replace Delta as the most prevalent SARS-CoV-2 variant across the world. Cryo-EM structural analysis of the Omicron variant spike protein in complex with human ACE2 reveals new salt bridges and hydrogen bonds formed by mutated residues R493, S496 and R498 in the RBD with ACE2. These interactions appear to compensate for other Omicron mutations such as K417N known to reduce ACE2 binding affinity, resulting in similar biochemical ACE2 binding affinities for Delta and Omicron variants. Neutralization assays show that pseudoviruses displaying the Omicron spike protein exhibit increased antibody evasion. The increase in antibody evasion, together with retention of strong interactions at the ACE2 interface, thus represent important molecular features that likely contribute to the rapid spread of the Omicron variant.

The Omicron (B.1.1.529) variant of SARS-CoV-2 binds to the hACE2 receptor more strongly and escapes the antibody response: Insights from structural and simulation data.

As SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) continues to inflict chaos globally, a new variant officially known as B.1.1.529 was reported in South Africa and was found to harbor 30 mutations in the spike protein. It is too early to speculate on transmission and hospitalizations. Hence, more analyses are required, particularly to connect the genomic patterns to the phenotypic attributes to reveal the binding differences and antibody response for this variant, which can then be used for therapeutic interventions. Given the urgency of the required analysis and data on the B.1.1.529 variant, we have performed a detailed investigation to provide an understanding of the impact of these novel mutations on the structure, function, and binding of RBD to hACE2 and mAb to the NTD of the spike protein. The differences in the binding pattern between the wild type and B.1.1.529 variant complexes revealed that the key substitutions Asn417, Ser446, Arg493, and Arg498 in the B.1.1.529 RBD caused additional interactions with hACE2 and the loss of key residues in the B.1.1.529 NTD resulted in decreased interactions with three CDR regions (1-3) in the mAb. Further investigation revealed that B.1.1.529 displayed a stable dynamic that follows a global stability trend. In addition, the dissociation constant (KD), hydrogen bonding analysis, and binding free energy calculations further validated the findings. Hydrogen bonding analysis demonstrated that significant hydrogen bonding reprogramming took place, which revealed key differences in the binding. The total binding free energy using MM/GBSA and MM/PBSA further validated the docking results and demonstrated significant variations in the binding. This study is the first to provide a basis for the higher infectivity of the new SARS-CoV-2 variants and provides a strong impetus for the development of novel drugs against them.

Insights into the specificity for the interaction of the promiscuous SARS-CoV-2 nucleocapsid protein N-terminal domain with deoxyribonucleic acids.

The SARS-CoV-2 nucleocapsid protein (N) is a multifunctional promiscuous nucleic acid-binding protein, which plays a major role in nucleocapsid assembly and discontinuous RNA transcription, facilitating the template switch of transcriptional regulatory sequences (TRS). Here, we dissect the structural features of the N protein N-terminal domain (N-NTD) and N-NTD plus the SR-rich motif (N-NTD-SR) upon binding to single and double-stranded TRS DNA, as well as their activities for dsTRS melting and TRS-induced liquid-liquid phase separation (LLPS). Our study gives insights on the specificity for N-NTD(-SR) interaction with TRS. We observed an approximation of the triple-thymidine (TTT) motif of the TRS to ß-sheet II, giving rise to an orientation difference of ~25° between dsTRS and non-specific sequence (dsNS). It led to a local unfavorable energetic contribution that might trigger the melting activity. The thermodynamic parameters of binding of ssTRSs and dsTRS suggested that the duplex dissociation of the dsTRS in the binding cleft is entropically favorable. We showed a preference for TRS in the formation of liquid condensates when compared to NS. Moreover, our results on DNA binding may serve as a starting point for the design of inhibitors, including aptamers, against N, a possible therapeutic target essential for the virus infectivity.

Discovery of Potential Antiviral Compounds against Hendra Virus by Targeting Its Receptor-Binding Protein (G) Using Computational Approaches.

Hendra virus (HeV) belongs to the paramyxoviridae family of viruses which is associated with the respiratory distress, neurological illness, and potential fatality of the affected individuals. So far, no competitive approved therapeutic substance is available for HeV. For that reason, the current research work was conducted to propose some novel compounds, by adopting a Computer Aided Drug Discovery approach, which could be used to combat HeV. The G attachment Glycoprotein (Ggp) of HeV was selected to achieve the primary objective of this study, as this protein makes the entry of HeV possible in the host cells. Briefly, a library of 6000 antiviral compounds was screened for potential drug-like properties, followed by the molecular docking of short-listed compounds with the Protein Data Bank (PDB) structure of Ggp. Docked complexes of top two hits, having maximum binding affinities with the active sites of Ggp, were further considered for molecular dynamic simulations of 200 ns to elucidate the results of molecular docking analysis. MD simulations and Molecular Mechanics Energies combined with the Generalized Born and Surface Area (MMGBSA) or Poisson-Boltzmann and Surface Area (MMPBSA) revealed that both docked complexes are stable in nature. Furthermore, the same methodology was used between lead compounds and HeV Ggp in complex with its functional receptor in human, Ephrin-B2. Surprisingly, no major differences were found in the results, which demonstrates that our identified compounds can also perform their action even when the Ggp is attached to the Ephrin-B2 ligand. Therefore, in light of all of these results, we strongly suggest that compounds (S)-5-(benzylcarbamoyl)-1-(2-(4-methyl-2-phenylpiperazin-1-yl)-2-oxoethyl)-6-oxo-3,6-dihydropyridin-1-ium-3-ide and 5-(cyclohexylcarbamoyl)-1-(2-((2-(3-fluorophenyl)-2-methylpropyl)amino)-2-oxoethyl)-6-oxo-3,6-dihydropyridin-1-ium-3-ide could be considered as potential therapeutic agents against HeV; however, further in vitro and in vivo experiments are required to validate this study.

Role of Q675H Mutation in Improving SARS-CoV-2 Spike Interaction with the Furin Binding Pocket.

Genotype screening was implemented in Italy and showed a significant prevalence of new SARS-CoV-2 mutants carrying Q675H mutation, near the furin cleavage site of spike protein. Currently, this mutation, which is expressed on different SARS-CoV-2 lineages circulating worldwide, has not been thoughtfully investigated. Therefore, we performed phylogenetic and biocomputational analysis to better understand SARS-CoV-2 Q675H mutants' evolutionary relationships with other circulating lineages and Q675H function in its molecular context. Our studies reveal that Q675H spike mutation is the result of parallel evolution because it arose independently in separate evolutionary clades. In silico data show that the Q675H mutation gives rise to a hydrogen-bonds network in the spike polar region. This results in an optimized directionality of arginine residues involved in interaction of spike with the furin binding pocket, thus improving proteolytic exposure of the viral protein. Furin was predicted to have a greater affinity for Q675H than Q675 substrate conformations. As a consequence, Q675H mutation could confer a fitness advantage to SARS-CoV-2 by promoting a more efficient viral entry. Interestingly, here we have shown that Q675H spike mutation is documented in all the VOCs. This finding highlights that VOCs are still evolving to enhance viral fitness and to adapt to the human host. At the same time, it may suggest Q675H spike mutation involvement in SARS-CoV-2 evolution.

Structure-Guided Design of Potent Inhibitors of SARS-CoV-2 3CL Protease: Structural, Biochemical, and Cell-Based Studies.

The COVID-19 pandemic is having a major impact on public health worldwide, and there is an urgent need for the creation of an armamentarium of effective therapeutics, including vaccines, biologics, and small-molecule therapeutics, to combat SARS-CoV-2 and emerging variants. Inspection of the virus life cycle reveals multiple viral- and host-based choke points that can be exploited to combat the virus. SARS-CoV-2 3C-like protease (3CLpro), an enzyme essential for viral replication, is an attractive target for therapeutic intervention, and the design of inhibitors of the protease may lead to the emergence of effective SARS-CoV-2-specific antivirals. We describe herein the results of our studies related to the application of X-ray crystallography, the Thorpe-Ingold effect, deuteration, and stereochemistry in the design of highly potent and nontoxic inhibitors of SARS-CoV-2 3CLpro.

Molecular Basis of SARS-CoV-2 Nsp1-Induced Immune Translational Shutdown as Revealed by All-Atom Simulations.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic represents the most severe global health crisis in modern human history. One of the major SARS-CoV-2 virulence factors is nonstructural protein 1 (Nsp1), which, outcompeting with the binding of host mRNA to the human ribosome, triggers a translation shutdown of the host immune system. Here, microsecond-long all-atom simulations of the C-terminal portion of the SARS-CoV-2/SARS-CoV Nsp1 in complex with the 40S ribosome disclose that SARS-CoV-2 Nsp1 has evolved from its SARS-CoV ortholog to more effectively hijack the ribosome by undergoing a critical switch of Q/E158 and E/Q159 residues that perfects Nsp1's interactions with the ribosome. Our outcomes offer a basis for understanding the sophisticated mechanisms underlying SARS-CoV-2 diversion and exploitation of human cell components to its deadly purposes.

Mechanistic insights into the effects of key mutations on SARS-CoV-2 RBD-ACE2 binding.

Some recent SARS-CoV-2 variants appear to have increased transmissibility compared to the original strain. An underlying mechanism could be the improved ability of the variants to bind receptors on the target cells and infect them. In this study, we provide atomic-level insights into the binding of the receptor binding domain (RBD) of the wild-type SARS-CoV-2 spike protein and its single (N501Y), double (E484Q, L452R) and triple (N501Y, E484Q, L452R) mutated variants to the human ACE2 receptor. Using extensive all-atom molecular dynamics simulations and advanced free energy calculations, we estimate the associated binding affinities and binding hotspots. We observe significant secondary structural changes in the RBD of the mutants, which lead to different binding affinities. We find higher binding affinities for the double (E484Q, L452R) and triple (N501Y, E484Q, L452R) mutated variants than for the wild type and the N501Y variant, which could contribute to the higher transmissibility of recent variants containing these mutations.

Porphyrin Molecules Decorated on Metal-Organic Frameworks for Multi-Functional Biomedical Applications.

Metal-organic frameworks (MOFs) have been widely used as porous nanomaterials for different applications ranging from industrial to biomedicals. An unpredictable one-pot method is introduced to synthesize NH2-MIL-53 assisted by high-gravity in a greener media for the first time. Then, porphyrins were deployed to adorn the surface of MOF to increase the sensitivity of the prepared nanocomposite to the genetic materials and in-situ cellular protein structures. The hydrogen bond formation between genetic domains and the porphyrin' nitrogen as well as the surface hydroxyl groups is equally probable and could be considered a milestone in chemical physics and physical chemistry for biomedical applications. In this context, the role of incorporating different forms of porphyrins, their relationship with the final surface morphology, and their drug/gene loading efficiency were investigated to provide a predictable pattern in regard to the previous works. The conceptual phenomenon was optimized to increase the interactions between the biomolecules and the substrate by reaching the limit of detection to 10 pM for the Anti-cas9 protein, 20 pM for the single-stranded DNA (ssDNA), below 10 pM for the single guide RNA (sgRNA) and also around 10 nM for recombinant SARS-CoV-2 spike antigen. Also, the MTT assay showed acceptable relative cell viability of more than 85% in most cases, even by increasing the dose of the prepared nanostructures.

Interaction of Spike protein and lipid membrane of SARS-CoV-2 with Ursodeoxycholic acid, an in-silico analysis.

Numerous repositioned drugs have been sought to decrease the severity of SARS-CoV-2 infection. It is known that among its physicochemical properties, Ursodeoxycholic Acid (UDCA) has a reduction in surface tension and cholesterol solubilization, it has also been used to treat cholesterol gallstones and viral hepatitis. In this study, molecular docking was performed with the SARS-CoV-2 Spike protein and UDCA. In order to confirm this interaction, we used Molecular Dynamics (MD) in "SARS-CoV-2 Spike protein-UDCA". Using another system, we also simulated MD with six UDCA residues around the Spike protein at random, naming this "SARS-CoV-2 Spike protein-6UDCA". Finally, we evaluated the possible interaction between UDCA and different types of membranes, considering the possible membrane conformation of SARS-CoV-2, this was named "SARS-CoV-2 membrane-UDCA". In the "SARS-CoV-2 Spike protein-UDCA", we found that UDCA exhibits affinity towards the central region of the Spike protein structure of - 386.35 kcal/mol, in a region with 3 alpha helices, which comprises residues from K986 to C1032 of each monomer. MD confirmed that UDCA remains attached and occasionally forms hydrogen bonds with residues R995 and T998. In the presence of UDCA, we observed that the distances between residues atoms OG1 and CG2 of T998 in the monomers A, B, and C in the prefusion state do not change and remain at 5.93 ± 0.62 and 7.78 ± 0.51 Å, respectively, compared to the post-fusion state. Next, in "SARS-CoV-2 Spike protein-6UDCA", the three UDCA showed affinity towards different regions of the Spike protein, but only one of them remained bound to the region between the region's heptad repeat 1 and heptad repeat 2 (HR1 and HR2) for 375 ps of the trajectory. The RMSD of monomer C was the smallest of the three monomers with a value of 2.89 ± 0.32, likewise, the smallest RMSF was also of the monomer C (2.25 ± 056). In addition, in the simulation of "SARS-CoV-2 membrane-UDCA", UDCA had a higher affinity toward the virion-like membrane; where three of the four residues remained attached once they were close (5 Å, to the centre of mass) to the membrane by 30 ns. However, only one of them remained attached to the plasma-like membrane and this was in a cluster of cholesterol molecules. We have shown that UDCA interacts in two distinct regions of Spike protein sequences. In addition, UDCA tends to stay bound to the membrane, which could potentially reduce the internalization of SARS-CoV-2 in the host cell.