Computational Analysis of SAM Analogs as Methyltransferase Inhibitors of nsp16/nsp10 Complex from SARS-CoV-2.

Methyltransferases (MTases) enzymes, responsible for RNA capping into severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), are emerging important targets for the design of new anti-SARS-CoV-2 agents. Here, analogs of S-adenosylmethionine (SAM), obtained from the bioisosteric substitution of the sulfonium and amino acid groups, were evaluated by rigorous computational modeling techniques such as molecular dynamics (MD) simulations followed by relative binding free analysis against nsp16/nsp10 complex from SARS-CoV-2. The most potent inhibitor (2a) shows the lowest binding free energy (-58.75 Kcal/mol) and more potency than Sinefungin (SFG) (-39.8 Kcal/mol), a pan-MTase inhibitor, which agrees with experimental observations. Besides, our results suggest that the total binding free energy of each evaluated SAM analog is driven by van der Waals interactions which can explain their poor cell permeability, as observed in experimental essays. Overall, we provide a structural and energetic analysis for the inhibition of the nsp16/nsp10 complex involving the evaluated SAM analogs as potential inhibitors.

Identifying inhibitors of NSP16-NSP10 of SARS-CoV-2 from large databases.

The COVID-19 pandemic, which has already claimed millions of lives, continues to pose a serious threat to human health, requiring the development of new effective drugs. Non-structural proteins of SARS-CoV-2 play an important role in viral replication and infection. Among them, NSP16 (non-structured protein 16) and its cofactor NSP10 (non-structured protein 10) perform C2'-O methylation at the 5' end of the viral RNA, which promotes efficient virus replication. Therefore, the NSP16-NSP10 complex becomes an attractive target for drug development. Using a multi-step virtual screening protocol which includes Lipinski's rule, docking, steered molecular dynamics and umbrella sampling, we searched for potential inhibitors from the PubChem and anti-HIV databases. It has been shown that CID 135566620 compound from PubChem is the best candidate with an inhibition constant in the sub-µM range. The Van der Waals interaction was found to be more important than the electrostatic interaction in the binding affinity of this compound to NSP16-NSP10. Further in vitro and in vivo studies are needed to test the activity of the identified compound against COVID-19.Communicated by Ramaswamy H. Sarma.

In silico study of potential antiviral activity of copper(II) complexes with non-steroidal anti-inflammatory drugs on various SARS-CoV-2 target proteins.

In silico molecular docking studies, in vitro toxicity and in silico predictions on the biological activity profile, pharmacokinetic properties, drug-likeness, ADMET (absorption, distribution, metabolism, excretion, and toxicity) physicochemical pharmacokinetic data, and target proteins and toxicity predictions were performed on six copper(II) complexes with the non-steroidal anti-inflammatory drugs ibuprofen, loxoprofen, fenoprofen and clonixin as ligands, in order to investigate the ability of these complexes to interact with the key therapeutic target proteins of SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) 3C-like cysteine main protease (3CLpro/Mpro), viral papain-like protease (PLpro), RNA-dependent RNA polymerase (RdRp), and non-structural proteins (Nsps) Nsp16-Nsp10 2'-O-methyltransferase complex, and their capacity to act as antiviral agents, contributing thus to understanding the role they can play in the context of coronavirus 2019 (COVID-19) pandemic. Cytotoxic activity against five human cancer and normal cell lines were also evaluated.

Exploring the Catalytic Mechanism of the RNA Cap Modification by nsp16-nsp10 Complex of SARS-CoV-2 through a QM/MM Approach.

The inhibition of key enzymes that may contain the viral replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have assumed central importance in drug discovery projects. Nonstructural proteins (nsps) are essential for RNA capping and coronavirus replication since it protects the virus from host innate immune restriction. In particular, nonstructural protein 16 (nsp16) in complex with nsp10 is a Cap-0 binding enzyme. The heterodimer formed by nsp16-nsp10 methylates the 5'-end of virally encoded mRNAs to mimic cellular mRNAs and thus it is one of the enzymes that is a potential target for antiviral therapy. In this study, we have evaluated the mechanism of the 2'-O methylation of the viral mRNA cap using hybrid quantum mechanics/molecular mechanics (QM/MM) approach. It was found that the calculated free energy barriers obtained at M062X/6-31+G(d,p) is in agreement with experimental observations. Overall, we provide a detailed molecular analysis of the catalytic mechanism involving the 2'-O methylation of the viral mRNA cap and, as expected, the results demonstrate that the TS stabilization is critical for the catalysis.

Identification of naphthyridine and quinoline derivatives as potential Nsp16-Nsp10 inhibitors: a pharmacoinformatics study.

This research is a recent effort to explore some new heterocyclic compounds as novel and potential nonstructural protein-16-nonstructural protein-10 (Nsp16-Nsp10) inhibitors for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) inhibition. The SARS-CoV-2 is causative agent of coronavirus disease 2019 (COVID-19) pandemic. A set of 58 molecules belongs to the naphthyridine and quinoline derivatives have been recently synthesized and considered for structure-based virtual screening against Nsp16-Nsp10. Molecular docking was virtually performed to screen for anti-SARS-CoV-2 activity against Nsp16-Nsp10. Fourteen out of fifty-eight compounds were exhibited binding affinity higher than co-crystal bound ligand s-adenosylmethionine (SAM) toward Nsp16-Nsp10. Further, the in silico pharmacokinetics assessment was carried out and it was found that two molecules possess the acceptable pharmacokinetic profile, hence considered promising Nsp16-Nsp10 inhibitors. The binding interaction analysis was revealed some crucial binding interactions between the final selected two molecules and ligand-binding amino acid residues of Nsp16-Nsp10 protein. In order to explore the characteristics of the protein-ligand complex and how selected small molecules retained inside the receptor cavity in dynamic states, all-atoms conventional molecular dynamics (MD) simulation was performed. Several factors were obtained from the MD simulation trajectory evidently suggested the potentiality of the molecules and stability of the protein-ligand complex. Finally, the binding affinity of both molecules and SAM was explored through the MM-GBSA approach which explained that both molecules possess strong affection towards the Nsp16-Nsp10. Hence, from the pharmacoinformatics assessment, it can be concluded that both heterocyclic compounds might be crucial for SARS-CoV-2 inhibition, subjected to experimental validation.Communicated by Ramaswamy H. Sarma.

Computational Investigation of Structural Dynamics of SARS-CoV-2 Methyltransferase-Stimulatory Factor Heterodimer nsp16/nsp10 Bound to the Cofactor SAM.

Recently, a highly contagious novel coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, has emerged, posing a global threat to public health. Identifying a potential target and developing vaccines or antiviral drugs is an urgent demand in the absence of approved therapeutic agents. The 5'-capping mechanism of eukaryotic mRNA and some viruses such as coronaviruses (CoVs) are essential for maintaining the RNA stability and protein translation in the virus. SARS-CoV-2 encodes S-adenosyl-L-methionine (SAM) dependent methyltransferase (MTase) enzyme characterized by nsp16 (2'-O-MTase) for generating the capped structure. The present study highlights the binding mechanism of nsp16 and nsp10 to identify the role of nsp10 in MTase activity. Furthermore, we investigated the conformational dynamics and energetics behind the binding of SAM to nsp16 and nsp16/nsp10 heterodimer by employing molecular dynamics simulations in conjunction with the Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PBSA) method. We observed from our simulations that the presence of nsp10 increases the favorable van der Waals and electrostatic interactions between SAM and nsp16. Thus, nsp10 acts as a stimulator for the strong binding of SAM to nsp16. The hydrophobic interactions were predominately identified for the nsp16-nsp10 interactions. Also, the stable hydrogen bonds between Ala83 (nsp16) and Tyr96 (nsp10), and between Gln87 (nsp16) and Leu45 (nsp10) play a vital role in the dimerization of nsp16 and nsp10. Besides, Computational Alanine Scanning (CAS) mutagenesis was performed, which revealed hotspot mutants, namely I40A, V104A, and R86A for the dimer association. Hence, the dimer interface of nsp16/nsp10 could also be a potential target in retarding the 2'-O-MTase activity in SARS-CoV-2. Overall, our study provides a comprehensive understanding of the dynamic and thermodynamic process of binding nsp16 and nsp10 that will contribute to the novel design of peptide inhibitors based on nsp16.

Structural insight into the recognition of S-adenosyl-L-homocysteine and sinefungin in SARS-CoV-2 Nsp16/Nsp10 RNA cap 2'-O-Methyltransferase.

The recent ongoing coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to rapidly spread across the world. To date, neither a specific antiviral drug nor a clinically effective vaccine is available. Among the 15 viral non-structural proteins (nsps), nsp16 methyltransferase has been considered as a potential target due to its crucial role in RNA cap 2'-O-methylation process, preventing the virus detection by cell innate immunity mechanisms. In the present study, molecular recognition between the two natural nucleoside analogs (S-adenosyl-l-homocysteine (SAH) and sinefungin (SFG)) and the SARS-CoV-2 nsp16/nsp10/m7GpppAC5 was studied using all-atom molecular dynamics simulations and free energy calculations based on MM/GBSA and WaterSwap approaches. The binding affinity and the number of hot-spot residues, atomic contacts, and H-bond formations of SFG/nsp16 complex were distinctly higher than those of SAH/nsp16 system, consistent with the lower water accessibility at the enzyme active site. Notably, only SFG could electrostatically interact with the 2'-OH and N3 of RNA's adenosine moiety, mimicking the methyl transfer reaction of S-adenosyl-l-methionine substrate. The atomistic binding mechanism obtained from this work paves the way for further optimizations and designs of more specific SARS-CoV-2 nsp16 inhibitors in the fight against COVID-19.