P.V. Krishna, M.R. Babu, and E. Ariwa (Eds.): ObCom 2011, Part I, CCIS 269, pp. 44–50, 2012. © Springer-Verlag Berlin Heidelberg 2012

Protein Ligand Interaction Studies of DJ-1 Protein Responsible for Parkinson’s Disease and Chemical

Analogues of Bacopa monnieri (Brahmi) Compounds

Subrata Sinha, Surabhi Johari, Khushboo Gupta, and Sewali Bora

Center for Bioinformatics Studies, Dibrugarh University, Dibrugah, Assam surabhibiotech1@gmail.com

Abstract. Parkinson’s disease (PD) is a neurodegenerative disorder characte-rized by progressive loss of dopaminergic neurons of substantia nigra . It has been discovered that DJ1 is a protein responsible for oxidative stress which causes aggregation of α synuclein fibrils further resulting in formation of Lewy bodies. These Lewy bodies serve as a marker for neuronal degeneration . There-fore probably inhibiting the DJ1 may reduce the formation of Lewy bodies thus decreasing the epidemiology of PD. Bacopa monnieri has been in use since early as a brain tonic to enhance memory development, however how the com-ponents of Bacopa monnieri functions is yet unknown. In this paper an attempt has been made to establish the component’s activities insilico with DJ1 that is responsible for Lewy bodies’ formation. We found that among the screened seven components, Jujubogenin glycosides interacts potentially with DJ1 but its drug like behavior was not satisfied so an analogue of Jujubogenin glyco-sides was designed which has passed all the properties of drug with drug likeliness 2.89 , drug score 0.65, highly soluble, highly permeable, non mutagenic, non tumerogenic and non irritant.

Keywords: Parkinson’s disease, DJ1, Bacopa monnieri, Analogue, Jujubogenin glycosides.

1 Introduction

A genetic study suggests that physiological effects of PD is in degeneration of a part of the brain called substania nigra ,through the loss of dopaminergic neurons which produce hormone dopamine [1]. The research is carried out in mice and drosophila to investigate protein DJ1 and its interactions with other genes [ 2]. It is known that mutations in the DJ-1 gene cause early onset autosomal receesive PD [3]. It was found that DJ1 is involved to promote neuron survival in aging cells which help us to study the cause of PD and its possible drugs [2].

The KEGG pathway entry hsa05012 also shows that the DJ1 protein is responsible for oxidative stress which causes aggregation of α Syn fibrils [4]. This aggregation of fibrils form lewy bodies which is considered as marker for neuronal degeneration

Protein Ligand Interaction Studies of DJ-1 Protein 45

[5].Thus inhibiting DJ1 protein may also inhibit formation of lewy body which further affects disease development and spread through the brain region.

At present there is no cure for Pakinson’s disease but a variety of medications pro-vides relief from the symptoms [6]. Usually patients are given levodopa combined with carbidopa. Nerve cells can use levodopa to make dopamine and replenish the brain’s dwindling supply [6] .Other drugs such as bromocriptine, pramipexole and ropinirole mimic the role of dopamine in the brain, causing the neurons to react as they would to dopamine [7]. An antiviral drug amantdine also appears to reduce symptoms [4].

In India for PD most of the time herbal medications “Bacopa Monnieri” also re-ferred as “Brahmi” has been used in the Ayurvedic system of medium centuries [8]. Traditionally it was used as a brain tonic to enhance memory development learning and concentration and provide relief to patients with anxiety or epileptic disorder [8]. .Bacopa herb contains many compounds including – Bacopaside I Bacopaside II, , Bacopaside III, Bacopaside IV, Bacopaside V, Bacopaside X, Bacopaside N2, Baco-pasaponin F, Bacopasoponin E, Bacopaside N1, Luteolin, Apigenin, Butalinic acid, Wogonin , Oroxindin, Pseudojujubogenin, Jujubogenin glycosides which is supposed to have ability to increase mental ability [9]. However how the components of Bacopa Monnieri interact with the various proteins of PD is yet unknown. Therefore in this paper an attempt has been made to establish drug likeliness of the various components of Bacopa monnieri insilico, by interaction studies of the components with the DJ1 one way responsible for PD.

2 Materials and Methods

2.1 Protein and Ligand Preparation

Referring the various literatures and KEGG pathway database [4] we found that prob-ably inhibition of DJ1 may effectively reduce the PD symptoms .The PARK 7 gene (accession number 11315) [10] on chromosome location 1p36.23 which encodes the protein DJ 1 was observed and retrived from Protein Data Bank (PDB) (http://www.rcsb.org/pdb/) with PDB id 3EZG [11] . The active sites of protein were identified by Molegro Virtual Docker [12&13].

The 3D structure of components of Bacopa monnieri and its physiochemical prop-erties were retrived from Chemspider Database for identifying its drug likeliness [14]. The components were filtered on the basis of Lipsinki’s rule of five using OSIRIS ((http://www.organicchemistry.org/prog/peo/) [15] and Molinspiration [16] that satis-fies Structure Activity Relationship (SAR) properties.

2.2 Optimisation of Protein and Ligand

The protein DJ downloaded from PDB (PDB id 3EZG) was prepared for docking by deleting all hetero atoms, ligands and water molecules and optimized by minimization

46 S. Sinha et al.

of energy by using AutoDock Vina [17] .The grid parameter was set and the obtained structure was saved as .pdbqt. Ligands- Ligands designed was obtained from Chemspider as mol format and was optimized by using AutoDock Vina. Later, all the optimized ligands were saved in .pdbqt format.The analogs were designed using Mer-vin sketch [18,19&20 ].

2.3 Molecular Descriptors

The molecular descriptors of screened five components of Bacopa monnieri were pre-dicted by loading them into an online server, OSIRIS property explorer [15]. This pre-diction process depends on comparison between precomputed set of structural moieties whose properties are already known and the structural moieties of loaded molecules. Molecular descriptors like clogP, solubility, drug score and side effects such as mutage-nicity, carcinogenicity and teratogenicity were determined. To calculate the overall drug score, OSIRIS combined logP, logS, molar mass, drug-likeness and toxicity risks into a single number to predict the molecule’s over all drug potential.

2.4 Docking Studies

All the 7 ligands , including Levodopa and were docked with DJ1 protein in the software AutoDock Vina (MGL Tools for Ligand Docking) [17] using a genetic algorithm and simulated annealing approach to explore wide range of ligand confor-mational flexibility and rotational flexibility of selected receptor hydrogens . In the aspect of producing binding energy estimates, the AutoDock Vina computes mini-mum binding energy with Jujubogenin glycosides as compared to other ligands [6]. The compound binding site was defined within a 10 Å radius around the binding sites of DJ1. Standard default parameter settings were used to evaluate the protein-ligand interactions [ 6 ].

2.5 Visualization of Docked Complex

The docked complex was visualized in Pymol [21] showing how the ligand interacts with the protein DJ1.

3 Results and Discussion

Target DJ1 (Pdb id 3EZG ) was downloaded from Protein Data Bank (PDB) Fig 1 .It is RNA binding protein consisting of A chains having molecular weight 20881.20 belonging to class alpha beta and DJ-1_Pfp protein family (PFam Accession no PF01965).

Protein Ligand Interaction Studies of DJ-1 Protein 47

Table 1. Toxicity Property of Ligands : OSIRIS Property Explorer

Ligand Mutagenic Tumorogenic Irritant Reproductive ClogP Solubility Molecular weight Drug likeness DrugScore

Wogonin No No No No 1.38 -2.85 284.0 1.71 0.83

Levodopa No No No No 0.94 -1.26 198 1.78 0.9

Luteolin Yes Yes No Yes 2.4 -2.56 286.0 1.9 0.32

Oroxondin Yes No No No -1.03 -2.48 464 0.44 0.5

Apigenin No No No Yes 2.42 -2.13 272 0.52 0.44

Jujubogenin – No No No No 5.05 -5.64 472 0.37 0.32

glycosides (JG)

Analogue of(JG) No No No No 2.97 -4.52 376 2.08 0.65

It was screened for its active site cavities by using by Molegro virtual docker. There was 1 predicted active site with site volume 255 Cubic Angstroms and Protein Volume: 17228 Cu-bic Angstroms. The cavity point was 30, - 11, 9 in x,y,z coordi-nates (Fig 2). DJ1 protein down-loaded from RCSB PDB server is prepared by removing water and other hetero molecules and it was geometrically optimized by Au-todock Vina. All the ligands are optimized to its least possible energy conformation. All the components of Bacopa monnieri were screened for its toxicity properties.

Among them the best compo-nents showing maximum drug likeliness are listed in Table 1 and Graph 1 where we found that Jujubogenin glycosides (JG) is the most important component of

Graph 1: Drug Likeliness Graph

Graph 2 : Solubility Pattern Graph

48 S. Sinha et al.

Bacopa monnieri having the properties non mutagenic,non tumerogenic ,non irritant and no reproductive effect and highly permeable (clogP=5.05) ,very soluble (-5.64) as per the range of clogP(<2.5) [23 ] and solubility (<1) [22] as shown in Graph 2 and 3 .Whereas the other components of Bacopa monnieri such as Lute-olin was (mutagenic and tumero-genic, drugscore=0.32), Oroxandin (mutagenic ,drug score=0.5) and Apigenin (reproductive effect , drug score=0.44) were found to be bad drug candidates It was found that JG inspite of following lipsnski rule of five but was not able to form docked complex with DJ 1 protein . So efforts were made to design analog of JG (Fig 3). The analogue showed maximum binding affinity with DJ-1 protein (Fig 4).

.

Fig. 1. The DJ1 Protein (PDB id 3EZG) obtained from Protein data bank

Fig. 2. Predicted Active Sites by Molegro Virtual docker visualized by PyMol. (Green)

No literature references described about the drug candidature of these components. It is assumed that analog of JG is having much higher external hydrogen bonding energy that leads to better binding than any other ligands. JG also shows good drug score when compared to other ligands as it shows higher H-bond & Vander Waal’s interactions.

Graph 3: Lipophilicity Graph Pattern

Protein Ligand Interaction Studies of DJ-1 Protein 49

Fig. 3. Analogue of Jujubogenin glyco-sides (JG) designed by Mervin Sketch

Fig. 4. The ligand (JG) inside the cavity of protein DJ 1

4 Conclusion

The interactions between DJ1 protein and the ligands were studied by using various computational methods. Based on binding energy, and hydrogen bond formed, dock-ing results were analyzed. The results were compared to find out the best ligand which can inhibit the property of the DJ1. Based on these observations, analog of JG is the most potent ligand among the other ligands . Thus the in silico method adopted in the present study helped in identifying the ligands using the commercial software and online tools for the treatment of neurodegenerative Parkinson’s disorder. This method reduces the time and cost in designing a drug as well as in analyzing the drug likeliness before it enters the clinical trials.

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