key: cord-0934004-3i7jdufv
authors: Rauf, Abdur; Rashid, Umer; Khalil, Anees Ahmed; Khan, Shahid Ali; Anwar, Sirajudheen; Alafnan, Ahmed; Alamri, Abdulhakeem; Rengasamy, Kannan RR
title: Docking-based virtual screening and identification of potential COVID-19 main protease inhibitors from brown algae
date: 2021-07-01
journal: S Afr J Bot
DOI: 10.1016/j.sajb.2021.06.033
sha: 3a5b39a72f10832e2e626593ce873be06d3154be
doc_id: 934004
cord_uid: 3i7jdufv

COVID-19 (SARS-CoV-2) is a viral disease that causes acute respiratory syndrome, which has increased the morbidity and mortality rate throughout the world. World Health Organization has declared this COVID-19 outbreak as pandemic and classified health emergency throughout the world. In the recent past, outbreaks of SARS and MERS have shown the interspecies transmission potential of coronaviruses and limitations of already prescribed drugs to overcome this global public health issue. Therefore, there is a dire need to identify a new regimen of targeted drugs from natural compounds having anti-COVID19 potential. This study aimed at screening 1018 brown algal natural compounds (many of them previously reported to have immunomodulatory effects) having probable anti-COVID19 potentials. The source compounds were extracted from MarinLit, a database dedicated to marine natural products and screened against COVID-19 main protease. The top seven compounds were further analysed, and their interactions with the active site were visualized. This study will further warrant screening the potent compounds against the virus in in-vitro conditions.

Since the very recent outbreak of SARS-CoV-2 (novel COVID-19) that has emerged as a global pandemic and public health emergency of international concern (Chow et al., 2020) , scientists worldwide have been working relentlessly to develop antivirals that can provide immunization against this deadly virus. Several studies have also been conducted to understand the virulence mechanisms of SARS-CoV-2. For instance, it has been suggested that SARS-CoV-2 can recognize human ACE2 more efficiently than the former SARS-CoV, thus increasing the capacity of the human-to-human transmission of SARS-CoV-2.

Furthermore, the SARS-CoV-2 spike protein can be seen to have a strong binding affinity with human ACE2. Consequently, the spike protein directly binds with the host cell surface ACE2 receptor, facilitating the virus entry and replication (Wan et al., 2020; Zhang et al., 2020c) . Hence, the ACE2 receptor can be regarded as an essential target against SARS-CoV-2.

Another desirable target for anti-corona virus drug design is the main protease which plays an essential role in viral gene expression and replication through the proteolytic processing of replicase polyproteins translated from the viral RNA (Xue et al., 2008; Zhang et al., 2020b) . Interestingly, the PDB 6LU7 protein structure, the main protease of SARS-CoV-2, has recently been determined (Liu et al., 2020) . Thus, the screening of potential protease inhibitors blocking the replication of coronavirus is of great relevance for developing anti-corona viral therapeutics.

With the urging need to search for safe and efficient antivirals against SARS-CoV-2 (COVID-19), natural products and, in particular, plants have been of considerable interest among researchers Mohammadi and Shaghaghi, 2020; Yang et al., 2020; Zhang et al., 2020a) . Most recently, a study by Thuy et al. (Thuy et al., 2020) showed that the compounds of garlic essential oil possessed anticoronavirus properties, as they were found to have good inhibitory activities on ACE2 and PDB6LU7 proteins.

In the last few decades, scientists have investigated a diverse class of natural products due to their pharmacologically active components. These natural products possess pharmacological properties having significant applications in the identification and isolation of bioactive compounds. Nearly 30% of biologically active formulations employed for the curtailment of various ailments are being extracted and isolated from natural products (Brito da Matta et al., 2011) . Among these, marine-derived natural compounds possess a wide range of new substances with various pharmacological potentials (Arif et al., 2004) . Since the last five decades, numerous investigations have demonstrated that marine algae (seaweeds), microorganisms, and marine invertebrates possess active compounds having therapeutic potentials against several diseases (Yasuhara-Bell et al., 2010) . The marine environment is considered to provide chemical diversity in the development of novel pharmacologically active agents. Additionally, seaweeds possess certain novel bioactive constituents having pharmacological activities, which are not present in terrestrial organisms (Gustafson et al., 2004) . These unique metabolites present in marine algae are phlorotannins, fucoidans, fucoxanthin, oxylipins, meroditerpenoids, and diterpenes (Glombitza and Gerstberger, 1985; Durán et al., 1997; Reddy and Urban, 2009; Biris-Dorhoi et al., 2020) . Various pharmacological application (antibacterial, antifungal, anti-inflammatory, neuroprotective, anti-tumor, etc.) of meroditerpenoids (Fallahydroquinone, Methoxybifurcarenone, & Sargaquinoic acid), phlorotannins (7,2''-Bieckol, 7-Hydroxyeckol hepta-acetate), and diterpenes (Triacetoxy-18-hydroxy-2,7-dolabelladiene) isolated from different species of marine brown algae have been reported in the literature (Bennamara et al., 1999; Horie et al., 2008; Kim et al., 2017; Ioannou et al., 2017; Hannan et al., 2020) .

Seaweeds are broadly classified into three classes, i.e. red seaweed (Rhodophyceae), green seaweed (Chlorophyceae), and brown seaweed (Phaeophyceae). These seaweeds are rich source of bioactive dietary components like proteins, polyunsaturated fatty acids, lipids, vitamins, terpenes (chromequinolide), dietary fibers, pigments (fucoxanthin, phycoerythrin, phycocyanin), colloids, minerals (Potassium, Iron, sulfur), aromatics, carbohydrates (fucoidan), acetogenins, polyphenols (phlorotannins), and antioxidants (de Souza Barros et al., 2015; Khalid et al., 2018) . Earlier, seaweeds were used by food processing and pharmaceutical industries due to their gelling properties, but now scientists are investigating their therapeutic role as complementary and alternative medicine. Various studies have highlighted the use of seaweeds to manage different diseases like cancer, obesity, diabetes, hypertension, inflammation, neurological disorders, viral and bacterial infections (Khalid et al., 2018) . Diverse class of primary and secondary natural compounds isolated from seaweed have shown promising effects against different viral, bacterial and fungal pathogens (Bedoux et al., 2017; Lima-Filho et al., 2002; Wang et al., 2012) . Initially, screening these natural compounds with pharmacological properties is necessary to evaluate them as potential inhibitors for these viral proteins. Therefore, keeping in view the emerging use of seaweed natural products and their pharmacological role, this study was designed in which a database of 1018 natural compounds from Brown algae was scanned against the covid-19 virus. The top seven compounds were analysed, and their interactions with the active site were visualized, which are presented herein.

For the Covid-19 main protease, crystal structure from Protein Data Bank (PDB ID = 5R82) in complex with 6-(ethylamino) pyridine-3-carbonitrile (RZS) was used as a template for the virtual screening (VS). The retrieved 3D crystal structure was prepared for the docking simulation by removing all the water molecules and any co-crystallized hetero molecules. For all atoms, the 3D-protonation was achieved in an implicit solvated environment (Temperature: 300 K; salt concentration: 0.1; pH: 7). 3D protonation was done for all atoms in an implicit solvated environment at pH 7, a temperature of 300 K, and a salt concentration of 0.1. The complete structure was energy minimized using Amber10:EHT force field until a root-mean-square (RMS) gradient of 0.1 kcal/mol/Å2 was reached. The centroid of the native ligand determined the active sites of the proteins.

One thousand eighteen (1018) brown algal compounds were extracted from the MarinLit database and used to test against the main protease receptor. Ligand.mdb database of compounds was built from SMILES format. By using the Amber10:EHT force field, the energy of compounds was minimized up to 0.001. The structure of the enzyme was opened in the Molecular Operating Environment (MOE) software.

The targeted Covid-19 main protease was subjected to docking-based virtual screening using the MOE dock program. The docking procedure was validated by re-docking of the native ligands. Comprehensive re-docking protocols were carried out to validate the docking algorithm. Native co-crystallized ligands were extracted and prepared comparably as for others. Docking was carried out using the Triangle matcher algorithm (placement stage) and scored by the London dG scoring function. Subsequently, best-scored poses were submitted to rigid receptor protocol (refinement stage). The final score was calculated with GBVI/WAS dGSF scoring function. Next, we changed the placement parameter from triangle matcher to alpha triangle and none. While rescoring was carried out using other two scoring functions (ASE and affinity dG).

To validate whether our approach can distinguish between active and inactive compounds, we have chosen randomly 1500 drug-like decoys from our in-house database of decoys [25] . We docked the 'validation set or decoy set' into the binding site of Covid-19 main protease (PDB ID = 5R82) as a positive control.

Docking simulation is considered an essential tool in drug discovery and the first step in any drug design process. The binding energy and the explanation of the protein inhibitor interactions are valuable results for further experimental and theoretical studies. The scores ranked the compounds by GBVI/WAS binding free energy calculation (in Kcal/mol). Compounds with binding energy values less than -8.500 kcal/mol were considered best for Covid-19 main protease inhibition. The top seven structurally diverse scaffolds were selected for further analysis ( Table 1) . All the selected compounds have shown the potential to inhibit the Covid-19 main protease. We also performed docking studies on lopinavir and Remdesivir as comparative standards. Moreover, we calculated the binding energy of phytochemical rutin found as the promising inhibitor of SARS-CoV-2 Mpro (Al-Zahrani 2020). The calculated binding energy values are enlisted in Table 2 . Met165 forms a π-sulfur interaction with a methoxy group. While the π-σ type of interactions with Thr25 also stabilizes the ligand-enzyme complex (Figure 2b) . 5-Hydroxy-cystofuranoquinol forms three hydrogen bond interactions with Thr24, Thr26, Ser46. Met165 forms πsulfur interactions with furan oxygen (Figure 3a) . Sargaquinoic acid forms three hydrogen bond interactions with Pro168, Gly143, and Gln189. Met165 forms a π-sulfur interaction with the methoxy group (Figure 3b) . showed better binding affinity (lowest binding energy) than lopinavir and Remdesivir used as comparative standards. Moreover, compounds also showed a better affinity with the enzyme than rutin.

The lack of specific COVID-19 drugs and/or vaccines and their rapid transmission rate has prompted scientists to investigate the new regimen of natural compounds having anti-COVID-19 potentials. Compounds from natural sources like seaweeds have shown their potential to combat various pathogenesis. Pathogenic SARS-CoV-2 has posed a significant threat to humankind in the last year or so, and therefore, the development of efficient therapeutics needs time. In the last decade, bioinformatics has played an essential role in rational drug discovery. In this study, a database of 1018 natural compounds from Brown algae was scanned against the COVID-19 virus. The top seven compounds were further analyzed, and their interactions with the active site were visualized. Molecular docking conducted in this study infers that all the compounds had significant binding in COVID-19 main protease (PDB ID = 5R82). Among all these examined compounds, 7,2''-Bieckol showed the lowest binding energy (high binding affinity, -10.7855 kcal/mol). Other compounds showed binding energy values in the range of -9.9611 to -8.5488 kcal/mol.

Considering that no specific drug has been formulated for COVID-19 infection, ligands identified in this study may help provide baseline data in developing multi-targeted drug discovery against SARS-CoV-2. On the other hand, further in vitro and in vivo studies must be conducted to authenticate the anti-COVID-19 potential of these identified compounds.

Funding: Abdulhakeem supported project number (TURSP-2020/288), Taif University, Taif, Saudi Arabia.

Rutin as a Promising Inhibitor of Main Protease and Other Protein Targets of COVID-19: In Silico Study

Novel marine compounds: anticancer or genotoxic?

Potential SARS-CoV-2 main protease inhibitors

Antiviral and cytotoxic activities of polysaccharides extracted from four tropical seaweed species

Methoxybifurcarenone: an antifungal and antibacterial meroditerpenoid from the brown alga Cystoseira tamariscifolia

Macroalgae-A Sustainable Source of Chemical Compounds with Biological Activities

Antinociceptive and anti-inflammatory activity from algae of the genus Caulerpa

Preliminary estimates of the prevalence of selected underlying health conditions among patients with coronavirus disease 2019-United States

Seaweeds with anti-herpes simplex virus type 1 activity

New diterpenoids from the alga Dictyota dichotoma

Putative inhibitors of SARS-CoV-2 main protease from a library of marine natural products: A virtual screening and molecular modeling study

Phlorotannins with dibenzodioxin structural elements from the brown alga Eisenia arborea

Antiviral marine natural products

Neuroprotective potentials of marine algae and their bioactive metabolites: Pharmacological insights and therapeutic advances

Antibacterial quinone metabolites from the brown alga, Sargassum sagamianum

Bioactive quinone derivatives from the marine brown alga Sargassum thunbergii induce anti-adipogenic and proosteoblastogenic activities

Dolabellanes with antibacterial activity from the brown alga Dilophus spiralis

Natural products and their derivatives against coronavirus: A review of the non-clinical and pre-clinical data

Therapeutic potential of seaweed bioactive compounds

Antibacterial activity of extracts of six macroalgae from the northeastern Brazilian coast

The crytal structure of 2019-nCoV main protease in complex with an inhibitor N3

Inhibitory effect of eight Secondary Metabolites from conventional Medicinal Plants on COVID_19 Virus Protease by Molecular Docking Analysis

The molecular docking study of potential drug candidates showing anti-COVID-19 activity by exploring of therapeutic targets of SARS-CoV-2

Meroditerpenoids from the southern Australian marine brown alga Sargassum fallax

Molecular docking analysis of selected natural products from plants for inhibition of SARS-CoV-2 main protease

Investigation into SARS-CoV-2 resistance of compounds in garlic essential oil

Natural and Nature-Derived Products Targeting Human Coronaviruses

Receptor recognition by the novel coronavirus from Wuhan: an analysis based on decade-long structural studies of SARS coronavirus

The antiviral activities and mechanisms of marine polysaccharides: an overview

Structures of two coronavirus main proteases: implications for substrate binding and antiviral drug design

Traditional Chinese medicine in the treatment of patients infected with 2019-new coronavirus (SARS-CoV-2): a review and perspective

In vitroevaluation of marine-microorganism extracts for anti-viral activity

Biologically active compounds from marine organisms in the strategies for combating coronaviruses

In silico screening of Chinese herbal medicines with the potential to directly inhibit 2019 novel coronavirus

Angiotensin-converting enzyme 2 (ACE2) as a SARS-CoV-2 receptor: molecular mechanisms and potential therapeutic target

Crystal structure of SARS-CoV-2 main protease provides a basis for design of improved α-ketoamide inhibitors

☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work