Larvicidal and antiviral nature of phoenix dactylifera L. natural products by targeting dengue virus and Aedes aegypti L. Proteins through molecular docking

1 Introduction

Dengue fever is a severe disease with a diagnostic rate of 2 million infections per year worldwide. One-third of the world's population is at risk of dengue virus infection. This disease is caused by the dengue virus, which spreads to people through the bites of infected Aedes species mosquitoes. DENV-1, DENV-2, DENV-3 and DENV-4 and DENV-5 are different dengue virus serotypes (Mustafa et al., 2015). They cause the same disease despite their genomic variation. A long single-stranded RNA encodes the polypeptide of the dengue virus. It is further cleaved into three structural proteins, including membrane, envelope, capsid, and seven non-structural (NS) proteins, including NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5(Endy et al., 2011; Normile, 2013; Qaddir et al., 2017).

Targeting DENV multifunctional enzymes are targeted to design antiviral therapies. NS2B/NS3 protease and NS3 helicase were targeted in this study. S2B-NS3 protease is the second-largest protein of the DENV genome (Chandramouli et al., 2010) and responsible for cytoplasmic cleavages, including at junctions between NS4B/NS5, NS3/NS4A, NS2B/NS3, and NS2A/NS2B protein and within the capsid, NS4A and NS2A proteins (Phoo et al., 2016). Similarly, NS3 helicase unwinds dsRNA to release ssRNA used as a template for NS5 protein in replication (Bartelma and Padmanabhan, 2002). NS2B/NS3 protease and NS3 helicase are conserved within the four serotypes (Chandramouli et al., 2010; Li et al., 2005), which permit the design of the drugs that are promising against the different serotypes (Keller et al., 2006; Xu et al., 2005).

Sterol carrier protein-2 (SCP-2) and kynurenine aminotransferase (KAT) are expressed throughout the animal kingdom, including insects. Aedes aegypti kynurenine aminotransferase (AeKAT) is a multifunctional enzyme that catalyzes the transamination of several amino acids and uses α-keto acids as amino group acceptors (Han et al., 2008). AeKAT has 45–50 % sequence identity with mammalian KAT-Is (Fang et al., 2002). AeKAT is mainly expressed in adult heads, demonstrating its significant role in the central nervous system (Fang et al., 2002). Aedes aegypti sterol carrier protein-2 protein (AeSCP-2) is an intracellular lipid carrier found in the midgut (Krebs and Lan, 2003). AeSCP-2 was reported to be involved in the cholesterol delivery and uptake across the cellular barrier between the hemocoel and the midgut (Blitzer et al., 2005; Dyer et al., 2003; Krebs and Lan, 2003).

Cholesterol is crucial for insects to grow, develop and reproduce (Perera and Wijerathna, 2019). AeSCP2 plays a vital role in cholesterol and fatty acid uptake in both larval and adult mosquitoes (Kumar et al., 2010). Knockdown of AeSCP-2 expression in larvae resulted in a reduction in cholesterol uptake, higher mortality, and decreased fecundity (Blitzer et al., 2005). In female adults, knockdown of AeSCP-2 expression leads to reduction of cholesterol uptake from the blood meal (Dyer et al., 2008). Few studies have focused on developing new mosquitocidal targeting AeSCP2 and AeKAT. Therefore, targeting AeSCP2 and AeKAT could be a substitute target for the discovery of promising mosquitocidal agents.

Medicinal plants have been used to treat various diseases since ancient times. Date palm, P. dactylifera L., a member of the Arecaceae family, is widely spread in the Arabian Peninsula. There are several potential health benefits of P. dactylifera (Al-Yahya, 1986; Allahyari et al., 2021; Demirci et al., 2013; Echegaray et al., 2020; Jassim and Naji, 2010). The crude extracts from this species have demonstrated antiviral (Allahyari et al., 2021) and insecticidal activities against Ae.aegypti (Demirci et al., 2011). A previous review on P. dactylifera reported more than 25 compounds, including phytoestrols, phytoestrogens, phenolic acid, and flavonoids (Al-Alawi et al., 2017). In the present study, these compounds will be used for docking against different target proteins of dengue virus and Ae. aegypti, and their interactions are observed to discover effective drugs.

Computational tools significantly impact drug discovery because of their fast and promising results. Docking studies predict the binding affinities of phytochemicals to the target enzymes. With our interest in searching for biological drug targets, we carried out this study to look for effective antiviral and mosquitocidal inhibitors in silico. The phytochemicals which show promising potential will be selected for further in vitro and in vivo testing in the future.

2 Materials and methods

3 Result

This evaluation of the bioactive compounds was based upon their binding parameters with the target proteins. The result of phytoestrols and phytoestrogens, phenolic acid, and flavonoid compounds against three different DENV enzymes (2FOM,3U1I, 2BMF) from Dengue Virus and three target proteins (1YIY, 1PZ4, 3UQI) from Ae. Aegypti (Table 1). The docked pose of 2FOM with the phytoesterol demonstrated the ligand's binding affinity with various targets of Dengue Virus (Table 2). The best potential binding sites of phytoestrols and phytoestrogens were −9.5 kcal/mol, −9.3 kcal/mol, and −8.9 kcal/mol for coumestrol, formononetin, and matairesinol, respectively. The best potential binding sites of phenolic acid and flavonoid compounds were 5-O-Caffeoylshikimic acid (-9.1 kcal/mol) and apigenin (-9.2 kcal/mol). In 2BMF, the potential binding sites among the different compounds docked were the best for pinoresinol (-9.5 kcal/mol) followed by luteolin (-9.0 kcal/mol) and chorogenic acid (-8.9 kcal/mol).

The target proteins of Ae. aegypti, namely 1YIY, 1PZ4, and 3UQI were docked with phytoestrols and phytoestrogens, phenolic acid, and flavonoid compounds by Autodock Vina. The energy values and the binding affinities are presented in Table 3. The energy values obtained of the drug targets of the most promising compound were recorded for 1YIY and 1PZ4 target proteins. The best potential binding sites of all compounds tested were recorded for isofucosterol (-10.3, kcal/mol) with 1YIY target proteins. This is followed by coumestrol (-9.3 kcal/mol) and campesterol (-8.9 kcal/mol) for the same target protein (1YIY). In 1PZ4, the best potential binding sites among the different compounds docked was isofucosterol (-9.7 kcal/mol) followed by coumestrol (-9.3 kcal/mol), p-fotmonrtin (-9.2 kcal/mol), and genistein (-8.9 kcal/mol). The best potential binding sites of phenolic acid and flavonoid compounds were apigenin (-9.2 kcal/mol) for 1YIY target proteins and chlorogenic acid (-8.6 kcal/mol) for 1PZ4 target proteins, respectively.

The docking poses and the 2D docking images of the best compounds with various Dengue virus and Ae. aegypti targets were analyzed with Protein-Ligand Interaction Profiler and PyMol software tools (Figs. 1-4). The docking poses were explored, and the amino acid residues involved in the various interactions were evaluated.

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Fig. 1

In silico molecular docking of the binding interaction of coumestrol compound with virus target protein (2FOM) based on the binding energy generated by AutoDock program. (A) A close-up view of the surface structure of 2FOM with coumestrol binding at the active site. (B) 2D structure of coumestrol interacting with 2FOM active site residues. …. Hydrophobic interaction Hydrogen bond.

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Fig. 2

In silico molecular docking of the binding interaction of pinoresinol compound with virus target protein (2BMF) based on the binding energy generated by AutoDock program. (A) A close-up view of the surface structure of 2BMF with pinoresinol binding at the active site. (B) 2D structure of pinoresinol interacting with 2BMF active site residues. …. Hydrophobic interaction Hydrogen bond.

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Fig. 3

In silico molecular docking of the binding interaction of isofucosterol compound with the mosquito target protein (1YIY) based on the binding energy generated by the AutoDock program. (A) A close-up view of the surface structure of 1YIY with isofucosterol binding at the active site. (B) 2D structure of isofucosterol interacting with 1YIY active site residues. - - - - - Hydrophobic interaction.

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Fig. 4

In silico molecular docking of the binding interaction of isofucosterol compound with mosquito target protein (1PZ4) based on the binding energy generated by AutoDock program. (A) A close-up view of the surface structure of 1PZ4with isofucosterol binding at the active site. (B) 2D structure of isofucosterol interacting with 1PZ4 active site residues. …. Hydrophobic interaction Hydrogen bond.

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The docking results of coumestrol to 2FOM resulted in four hydrogen bonds (ARG24, GLN25, VAL26) and ten hydrophobic interactions (ILE12, LEU16, ILE19, VAL26, LEU48, LEU102, PHE105, ILE106, LEU109) with binding energies of −9.5 kcal/mol. Similarly, the docking simulation of 2BMF to pinoresinol resulted in seven hydrogen bonds to THR289, ASP290, ARG322, ASP409, and GLU514, resulting in the energy of − 9.5 kcal/ mol. There were also two hydrophobic (THR289 and THR450) and one π-cation (LYS515) interaction with amino acid residues.

Among the 23 phytocompounds, isofucosterol exhibited the best binding conformations with the lowest binding energy values with the two target proteins 1YIY and 1PZ4 of Ae.egyptia. The docking simulation of 1PZ4 to isofucosterol resulted in one hydrogen bond (GLY17) with − 9.7 kcal/mol binding energies. Also, formation of 16 hydrophobic interactions with ILE12, ARG15, LEU16, ILE19, ASN23, ARG24,GLN25, VAL26, LEU102, PHE105, ILE106, and LEU109. A similar observation of target protein 1YIY was noted with isofucosterol, with –10.3 kcal/mol energy. No hydrogen bond interaction was observed with isofucosterol, but there were 13 hydrophobic interactions with TRP27, ILE31, GLN44, PHE46, PHE135, VAL223, TYR224, LYS255, and THR260.

4 Discussion

Computational advances have a significant influence on the process of drug development. Virtual screening is widely used to reduce the time and cost of drug discovery. Molecular docking is an approach used to discover novel ligands for a target protein and plays an essential role in structure-based drug design (Meng et al., 2011). The relationship between the receptor and ligand plays a vital role in drug formulations. Several drugs were isolated from the natural products based on bioactivity–guided fractionation. Natural products can help cure many human diseases.

Natural products-derived antiviral compounds such as alkaloids (Acridone, Aporphine, β-Carboltne), carbohydrates (Glucosamine, Sulphated polysaccharides, γ-carragenan), chromones (Khellin, Visnagin, Psoralen), furanocoumarins and flavonoids (Cyanidin, Pelargodin), phenolics (Benzoic acid and Caffeic acid derivatives), terpene (Scopadulcic acid β, Scopadulin), triterpene steroids (Nigranoic acid, Buxamine E, Cyclobuxamine H) and many more have been published in several review articles (Che, 1991; El Sayed, 2000; Meng et al., 2011; Rinehart et al, 1993). Similarly, Several compounds have been published dealing with natural products-derived larvicidal compounds such as octacosane isolated from Moschosma polystachyum, falcarinol and falcarindiol (Cryptotaenia Canadensis), trans-asarone (Daucus carota), 3-n-butyl-4,5- dihydrophthalide (Apium graveolens), geranial, and neral (Magnolia salicifolia), β-selinene (Apium graveolens), β-selinene (graveolens, neoduline), 4-methoxyneoduline, and nepseudin ((Neorautanenia mitis) and many more (Kishore et al., 2014).

This study investigated the binding capability of bioactive compounds from P. dactylifera with key antiviral and larvicidal protein targets since various biological activities had been reported from this plant. Our study shows that the selected compounds can efficiently bind to the target protein and that molecular docking can be successfully used to find inhibitors from P. dactylifera.

Dengue protease plays an important role in dengue virus replication (Geiss et al., 2009). Therefore, blocking the interaction between its subunits or the active site disturb viral replication in the host. Reports revealed that disulphide cyclic peptides have inhibition potential against dengue protease(Tambunan et al., 2011; Trott and Olson, 2010). Therefore, pinoresinol and coumestrol reported in this study may obstruct the activity of the targeted protein via affecting the interactions with the active site or blocking the binding of subunits necessary for complex formation.

Mosquito control is carried out either on adults or immature larvae. The use of synthetic insecticides in controlling mosquito's adults and the larval population is harmful to the environment. Therefore, this necessitates the search for eco-friendly mosquitocides for controlling the mosquitoes' larvae.

AeKATis a multifunctional aminotransferase that catalyzes the transamination of several amino acids and uses α-keto acids as amino group acceptors (Han et al., 2008). It has a vital role in neuroactive activity (Han and Li, 2004). AeKAT is mainly expressed in adult heads of mosquitoes, demonstrating its significant role in the CNS (Fang et al., 2002). Cholesterol uptake is essential for the larval population and is carried out through the AeSCP-2. Several compounds were screened to block the target protein. Currently, isofucosterol was a promising analog to dock with cholesterol carrier protein AeSCP-2.

Many studies have attempted to look for different sources of compounds that inhibit the action of SCP-2 via competitive binding. Some are natural compounds (Anstrom et al., 2012; Ee et al., 2006; Larson et al., 2014), and some are synthetic compounds (Kim et al., 2005),. Kim et al. reported several sterol carrier protein inhibitors. They found that these inhibitors showed physiological effects on cholesterol metabolism in cell culture, similar to the impact of AeSCP-2 knockdown (Kim et al., 2005). Several studies revealed that sterol carrier protein inhibitors have toxicity on Cx. pipiens, Cx. quinquefasciatus, and Anopheles gambiae (Larson et al., 2014; Li et al., 2009). Furthermore, it is reported that there is a synergistic activity of AeSCP-2 inhibitors when used with permethrin (Li et al., 2009). Furthermore, the AeSCP-2-mediated cholesterol uptake pathway is required for dengue virus production in Aedes mosquitoes, and the inhibition of AeSCP-2 activity resulted in repressed production of the virus within mosquito cells in vitro (Fu et al., 2015). However, further studies are required to elucidate the interaction mechanism between the promising compounds, the targeted proteins, and the feasibility of using these inhibitors in the fields.