Insilico Docking Studies of Some Isolated Selected Compounds of Phoenix sylvestris (L.) Against Thrombosis

Md Alamgir Hossain1, Md Towhidul Islam Tarun2, Md. Anwar Morshed3, Md Salman Faisal4, Whaheduzzaman5, Md Mostafa Moin Uddin6, Md Ershad1, Afzal Hosen1, Md AL Amin1 and Mohuya Majumder*7,8 1Department of Chemistry, Jahangirnagar University, Bangladesh 2Department of Pharmacy, Jahangirnagar University, Bangladesh 3Department of Pharmacy, International Islamic University Chittagong 4Department of Pharmacy, Manarat International University, Bangladesh 5Department of Pharmacy, Jagannath University, Bangladesh 6Department of Chemistry, Chittagong University, Bangladesh 7Department of Pharmacy, East West University, Bangladesh 8GUSTO A Research Group, Bangladesh


Introduction
Thrombosis is the formation of a blood clot inside a blood vessel, obstructing the flow of blood through the circulatory system. A thrombus is more likely to occur in people who are immobile, and who are genetically predisposed to blood clotting. It can also form if an artery, vein, or surrounding tissue is damaged. A thrombus can block the flow of blood through a vein or artery if it detaches from the vessel wall and lodges in the lungs or other vital organs, it can become a life-threatening embolus [1]. Thrombolysis is the breakdown (lysis) of blood clots by pharmacological means, and commonly called clot busting. It works by stimulating secondary fibrinolysis by plasmin through infusion of analogs of tissue plasminogen activator (tPA), the protein that normally activates plasmin. Thrombolysis mainly involves the use of thrombolytic drugs, which dissolve blood clots [2]. The purpose of a fibrinolytic drug is to dissolve thrombin in acutely occluded coronary arteries thereby to restore blood supply to ischemic myocardium, to limit necrosis and to improve prognosis [3]. For the treatment of myocardial infarction, many thrombolytic agents are used. Among them, streptokinase is remarkable and widely used. Moreover, Tissue-type Plasminogen activator is more effective and safer than either urokinase or streptokinase type activators. It is noted that all available thrombolytic agents still have significant deficiencies, including the necessity of large doses to be maximally effective, limited fibrin specificity and a significant associated bleeding tendency. Therefore, steps are taken to develop improved recombinant variants of these drugs in order to minimize deficiencies of the available thrombolytic drugs [4][5][6][7][8].
The trend of using natural products has increased together with active plant extracts are normally for new drug discoveries [9].
Plants have been the time frame of countless traditional medicine systems across the world for thousands of years and continue and offer people with brand-new cures. Phoenix sylvestris Roxb. (Arecaceae), locally known as Khejur, is a palm tree cultivated for its syrupy juice and edible fruit in Bangladesh [10]. Palm is one of the important horticultural crops in many countries [11]. In Bangladesh, Khejur palm is produced as a homestead crop; however, it grows naturally or is cultivated in fallow lands, around homesteads, farmland boundary and even in the marginal lands along the roads and canals [12]. Fruits of the plant are used to treat back pain, stomachache, toothache, headache, arthritis, pain of buttocks, fever, piles, nervous debility, and as nervine tonic, restorative, sedative in ethnomedicine [10,13]. In addition, it is widely used as an aphrodisiac, sweetener and diuretic and in the treatment of vomiting, vertigo and unconsciousness. It improves cardiovascular health by soaking out all the cholesterol from the arteries. They have high calcium content and improve bone health. Generally, the juice of P. sylvestris is consumed as a cooling beverage [14]. The sap of Khejur palm is a good source of vitamins of the B group and contains, in addition, a variable amount of ascorbic acid [15], freshly harvested sap consists of sucrose around 10%, minimal invert sugar of <0.5% and a small amount of protein, gums, and minerals.
Computational simulations of drug-target interactions using in silico molecular docking and molecular dynamics approaches are commonly used for the rational design and screening of drugs [16]. Molecular docking has become a major computational method for the prediction of ligand-receptor interactions [17]. A productive docking strategy must have the ability to adequately envision the local ligand represent the receptor limiting site (i.e. to find the trial ligand geometry inside a resistance confine) and the related physical-compound sub-molecular affiliations [18][19][20].
Furthermore, the aim of the study to find the mechanism of action of the isolated compounds from Phoenix sylvestris was explored the thrombolytic activity by molecular docking analysis.

In silico Molecular Docking Protein Preparation
Three-dimensional crystal structure of Catalytic domain of human two-chain tissue plasminogen activator complex of a bisbenzamidine (PDB id: 1A5H) was downloaded in pdb format from the protein data bank [21]. After that, the structure was prepared and refined using the Protein Preparation Wizard of Schrödinger-Maestro v10.1. Charges and bond orders were assigned, hydrogens were added to the heavy atoms, selenomethionines were converted to methionines and all waters were deleted. Using force field OPLS_2005, minimization was carried out setting maximum heavy atom RMSD (root-mean-square-deviation) to 0.30Å.

Ligand Preparation
The six major representative compound structures i.e., Diethyl

Receptor Grid Generation
Receptor grids were calculated for prepared proteins such that various ligand poses bind within the predicted active site during docking. In Glide, grids were generated keeping the default parameters of van der Waals scaling factor 1.00 and charge cut

Glide Standard Precision (SP) Ligand Docking
SP flexible ligand docking was carried out in Glide of Schrödinger-Maestro v 10.1 [22]. Within which penalties were applied to non-cis/trans amide bonds. Van der Waals scaling factor and partial charge cutoff were selected to be 0.80 and 0.15, respectively for ligand atoms. Final scoring was performed on energy-minimized poses and displayed as Glide score. The best-docked pose with lowest Glide score value was recorded for each ligand.

In silico Molecular docking analysis
Prediction of interaction energies between ligand and receptor has been a major challenge for molecular docking [23]. Virtual screening utilizes docking and scoring of each compound from a dataset. The technique used is based on predicting the binding modes and binding affinities of each compound in the dataset by means of docking to an X-ray crystallographic structure [24]. Gridbased docking study was used to analyze the binding modes of molecules with the amino acids present in the active pocket of the protein [25].To identify the potential thrombolytic lead molecule,  Table   1 and the docking figure showed in Figure1. The results of docking analysis were described in Table 1  Ligand atoms that are exposed to solvent are marked with gray spheres. The protein "pocket" is displayed with a line around the ligand, colored with the color of the nearest protein residue. The gap in the line shows the opening of the pocket.  Insilco model support that all the isolated compound from P.
sylvestris might be a TISSUE PLASMINOGEN ACTIVATOR inhibitor.
Further in vivo investigation needs to identify the potential inhibitory activity of isolated compounds from P. sylvestris.

Competing Interests
The authors declare that they have no competing interests.