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MOLECULAR DOCKING ANALYSIS OF QUERSETIN, GUAIJAVARIN AND

MORIN-3-O--L-ARABOPYRANOSIDE AS ANTIBACTERIAL AGENT

Raisa Fadilla1, Asmiyenti Djaliasrin2, Rina Herowati1


1
Faculty of Pharmacy, Setia Budi University, Jl. Letjen Sutoyo Solo 57127, Indonesia
2
Faculty of Pharmacy, Muhammadiyah University, Jl. Raya Dukuhwaluh Purwokerto

53182, Indonesia

Email:Raisa.fadilla@gmail.com

ABSTRACT

Guava leaves have been proven to have antibacterial activity and have been

widely used for the prevention and therapeutic treatment of diarrhea. Guava leaves

contain many compounds that have antibacterial activity. Some of these compounds

are quercetin, guaijavarin and morin-3-O--L-lyxopyranoside. This study aims to

understand the interaction and interaction patterns of quercetin, guaijavarin and morin-

3-O--L-arabopiranocid compounds of guava leaves (P. guajava L.) towards 9

biological molecule antibacterial targets through molecular docking. Biological

molecule antibacterial targets used were SleB protein (4F55), SleL protein (4S3J),

maltoporin (1MPR), Agga (aggregative adherence fimbriae / 1) (4PH8), Bc1960

peptidoglycan N-acetylglucosamine deacetylase (4L1G), phosphatidylcholine-

phospholipase C Bc (2HUC), phospholipase C regulator + PAPR (2QFC), penicillin

binding protein 3 E.coli (transferase) (4BJP) and gyrase + 4.5-dibromopyrolamide-

based inhibitor (isomerase) (4ZVI).

In this study, molecular docking performed on quercetin, guaijavarin and morin-

3-O--L-lyxopyranoside compounds of guava leaves (P. guajava L.) as ligand towards

9 biological molecule antibacterial targets using Autodock Vina software as well as

PyMOL as visualization tools. guaijavarin, morin-3-O--L-arabopiranocid and quercetin


compunds of guava leaves (P. guajava L.) interact well with 7 biological molecule

antibacterial targets because it can bind to the active site residues of the

macromolecular protein 4F55, 1MPR, 4L1G, 2HUC, 2QFC, 4ZVI and 4BJP with Gbind

range -6,5-9,6 kcal/mol and shows the interaction patterns that resemble the original

ligand to the protein macromolecules 2QFC and 4ZVI AS seen from visual observation.

Keywords: Antibacterial, molecule docking, quercetin, guaijavarin, morin-3-O--L-

lyxopyranoside

INTRODUCTION

Diarrhea is a disease indicated by increased frequency of defecation more than

usual (> 3 times/day) along with a change in feces consistency (to liquid), with / without

blood and / or mucus [1]. Bacteria that cause diarrhea can be divided into two major

categories, the bacteria non-invasive and invasive bacteria. Included in the class of

non-invasive bacteria are: V. cholerae, pathogenic E. coli (EPEC, ETEC, EIEC), while

groups of invasive bacteria are Salmonella sp [2]. Most microorganisms that cause

diarrhea spread via fecal-oral route through food, contaminated water or transmitted

between humans by close contact [3].

Diarrhea treatment which allegedly caused by invasive bacterial infections are

treated with antibiotics. Treatment with antibiotics is only needed on serious form of

diarrhea bacteria. The main choices are amoxicillin, cotrimoxazole and fluoroquinolone

compound. These drugs should not be given more than 7-10 days, except where after

recovering the diarrhea, the patient was still issuing the bacteria in the feces [4]. Giving

antimicrobials should consider the clinical benefits with cost, risk of side effects,

eradication of the normal flora intestinal that is harmful, induction of Shiga toxin

production and rising resistance to antimicrobial [5].


Traditional medicine is widely used by the society because it is natural, easy to

obtain, and the price is cheap, the use of traditional herbs does not create side effects,

as it often happens in the chemical treatment, other than that there are still many

people who believe that the use traditional medicine is safer than synthetic drugs [6].

Guava leaf (P.guajava L.) has been widely used by the public as a treatment of

diarrheal diseases. The compounds contained in guava leaves have antibacterial

activity based on the acquired MIC values. Some of these compounds include

quercetin, guaijavarin and morin-3-O--L-arabopiranocid [7].

This study aimed to analyze whether the quercetin, guaijavarin and morin-3-O-

-L-arabopiranocid compounds of guava leaves (P. guajava L.) has interactions to 9

biological molecule antibacterial target and predict interaction patterns of quercetin,

guaijavarin and morin-3-O--L-arabopiranocid compounds toward targets through the

molecular docking. Method used in the study is molecular docking. Docking is a

method that can predict the orientation of a molecule to another molecule when it binds

to form a stable complex [8]. Molecular docking method is a computing system on

biological screening, the goal is to find the value, rank or filter the set of data structures

using one or more procedures computing [9]. This method is useful to provide initial

knowledge about the type of drug compounds as ligand binding with specific

macromolecules [10].

EXPERIMENTAL SECTION

Materials

The materials used in this study were SleB protein (PDB ID: 4F55), SleL protein

(PDB ID: 4S3J), maltoporin (PDB ID: 1MPR), Agga (aggregative adherence fimbriae /

1) (PDB ID: 4PH8), Bc1960 peptidoglycan N-acetylglucosamine deacetylase (PDB ID:

4L1G), phosphatidylcholine-phospholipase CBC (PDB ID: 2HUC), phospholipase C


regulator + PAPR (PDB ID: 2QFC), penicillin binding protein 3 E.coli (transferase)

(PDB ID: 4BJP) and gyrase + 4.5-dibromopyrolamide-based inhibitor (isomerase) (PDB

ID: 4ZVI) downloaded from the protein Data Bank. The ligands used were quercetin,

guaijavarin and morin-3-O--L-arabopiranocid compounds drawn and performed

geometry optimization using ACD / ChemSketch. 3D structure can be viewed using

JMOL.

Instrumentation

The instruments used in this study were a set of PC Laptop Asus Intel Core i7-

4720 X450J HQ, 3.6 GHz, 4 GB RAM with Microsoft Windows XP. The software used

among others JMOL, ACD / ChemSketch, Discovery Studio, PyRx-Autodock-Vina,

PyMOL which supported with internet access to make connections with an online

program.

Procedure

Preparation of Molecular Structure Biology Antibacterial Targets

SleB protein (4F55), SleL protein (4S3J), maltoporin (1MPR), Agga

(aggregative adherence fimbriae / 1) (4PH8), Bc1960 peptidoglycan N-

acetylglucosamine deacetylase (4L1G), phosphatidylcholine-phospholipase CBC

(2HUC), phospholipase C regulator + PAPR (2QFC), penicillin binding protein 3 E.coli

(transferase) (4BJP) and gyrase + 4.5-dibromopyrolamide-based inhibitor (isomerase)

(4ZVI) were downloaded from the protein Data Bank. Furthermore these targets were

optimized with Autodock Tools saved with .pdbqt format. The conducted optimization

was the removal of water molecules and original ligand bound by using Discovery

Studio and the addition of a hydrogen atom.


Preparation of Ligand Structure

Quercetin, guaijavarin and morin-3-O--L-arabopiranocid were drawn and

performed geometry optimization using ACD / ChemSketch, the appearance of the 3D

structure could be viewed by using JMOL. Furthermore ligand format converted into

.pdbqt by using Discovery Studio. The conducted optimization was the addition of a

hydrogen atom in the ligand which will automatically increase when the Autodock Tools

program and setting number of activesite torsion opened.

Validation of Molecular Docking

Ligand docking validation and ligand was added to the original program-

Autodock PyRx-Vina. Selected ligand and macromolecular protein were used to run

docking process. Ligand resulted from docking process were saved (ligand validation)

and compared with original ligand to see the value Root Mean Square Deviation

(RMSD). Docking results might be continued if it has a value of 2 RMSD.

Molecular docking

Biological molecule antibacterial target and ligands of quercetin, guaijavarin

and morin-3-O--L-arabopiranocid which had been prepared incorporated into

Autodock PyRx-Vina program. The performed procedure was similar to the docking

validation; however comparison would not be done. The initial stage in conducting

docking with Autodock Vina was by opening the ligand file and macromolecular protein

after optimization. Then the grid box parameter settings, parameter box for ligand

2QFC grid was done and 4ZVI equated with grid box parameter of original ligand when

validation was conducted. Then the parameter of grid box test ligand controlled by

pointing to the active side of the macromolecules protein. After setting the grid box,

then continue with the process of docking.


Analysis and Visualization of molecular docking

The determination of ligand conformation docking result was done by selecting

ligand conformations which had the lowest binding free energy (best pose) and looking

at the pattern of formed interaction, the more similar the patterns of formed interaction

it means ligand had the same status as the original and complex ligand-protein became

more stable and ligan became more potent. The position and orientation of the ligands

on the macromolecules, as well as amino acids bounded to ligands were visualized by

software PyMOL.

RESULTS AND DISCUSSION

Validation was performed on the regulator phospholipase C + PAPR (2QFC)

and gyrase + 4.5-dibromopyrolamide-based inhibitor (isomerase) (4ZVI) using PyRx-

Autodock-Vina. The obtained docking results showed that the original tethered ligand

could return to the same position prior to separation of the ligand to the protein

macromolecules. It could be seen by looking at the amino acids that was bound or

nearby to a ligand. Amino acid protein macromolecules that were bound to a ligand

prior to separation could be seen through the binding pocket. Original ligands 2QFC

were ligan leu-pro-phe-glu-phe, on ligands leu-pro-phe-glu-phe could be seen ligand

bound with 163 Asn, 201 Asn, 204 Lys and 275 Tyr, beside that the other visible amino

acids were 200 Tyr, 167 Glu and 197 Lys. After being compared with the results of the

validation docking, there were similarities between the original ligand with the results of

the binding validation of the amino acid residues via second overlay ligands, which

hydrogen bonds were formed at Lys 204 and the same amino acid binding around

ligands, which were Tyr 200 and Glu 167 with RMSD <2 , which was 1.529 in the

center-x: -22.1, center-y: 52.6, center-z: -0.3, size-x: 17, 5, size-y: z size-13.2 and:

20.2. The original ligand was ligand 4S4 4ZVI could be seen hydrogen bond at amino

acid Asp 73 as well as the validation results were visible through the overlay both
ligands with RMSD <2 , which is 1.178 in the center-x: -14.1, center-y: 17.5, center

z: 25.1, size-x: 22.7, size-y: z size-20.5 and: 19.5. Overlay results between the original

ligand with ligand docking results showed that the original ligand with ligand docking

results had similar position and interaction patterns.

Based on the analysis of the obtained compiled data, it was suspected that

biological molecules targets of antibacterial proteins SleL (4S3J) and AggA

(aggregative adherence fimbriae / 1) (4PH8) was not a target of quercetin, guaijavarin

and morin-3-O--L lyxopyranoside compounds. Furthermore, judging from Gbind

and patterns of interaction that occurred, three best data could be seen, which are the

interaction of guaijavarin and quercetin compounds in molecular biology targets

antibacterial maltoporin (1MPR) and guaijavarin compound on the molecular biology

targets antibacterial phosphatidylcholine-phospholipase C Bc (2HUC) .

CONCLUSION

Quercetin, guaijavarin and morin-3-O--L- arabopiranocid compounds of guava

leaves (P. guajava L.) shows the interaction towards 7 biological molecule antibacterial

targets 4F55, 1MPR, 4L1G, 2HUC, 2QFC, 4ZVI and 4BJP and have interaction

patterns that resemble the original ligand towards a biological molecule antibacterial

targets 2QFC and 4ZVI when viewed from visual observations through molecular

docking.

REFERENCES

1. Suraatmaja, Sudaryat. 2007. Kapita Selekta Gastroentrologi Anak. Jakarta: CV

Sagung Seto.

2. Vila J, Vargas M, Ruiz J, Corachan M, De Anta MTJ, Gascon J. 2000: Quinolon

Resisten in Enterotoxigenic E.colli causing Diarrhea in Travelers to India in


Comparison with other Geographycal Areas. Antimicrobial Agents and

Chemotherapy.

3. Wong, L.D., Eaton, H.M., Wilson, D., Winkelstein, L.M., dan Schwart, P., 2009.

Buku Ajar Keperawatan Pediatrik. EGC. Jakarta.

4. Tjay, Tan Hoan dan Kirana Rahardja. 2008. Obat-Obat Penting Khasiat,

Penggunaan dan Efek-Efek Sampingnya Edisi Keenam. Jakarta: PT. Elex Media

Komputindo.

5. Eppy. 2009. Diare Akut. MEDICINUS Scientific Journal of Pharmaceutical

Development and Medical Application. 22(3), 91-98.

6. Thomas, A.N.S. 1989. Tanaman Obat Tradisional. Yogyakarta: Kanisius.

7. Arima H dan Gen-ichi D. 2002. Isolation of Antimicrobial Compounds from Guava

(Psidium guajava L.) and their structural elucidation. Bioscience, Biotechnology,

and Biochemistry. 66(8), 1727-1730.

8. Lengauer T, Rarey M. 1996. Computational methods for biomolecular docking.

Curr OpinStruct Biol. Jun;6(3):402-6. Review. PubMed PMID: 8804827.

9. Leach, A., Shoicet, B., & Peishoff, C. 2006. Docking and Scoring. J Med Chem 49

(20).

10. Klebe, G. 2005. Virtual Screening: Scope and Limitations. In J. Alvarez, & B.

Shoichet (Ed). Virtual Screening in Drug Discovery. Boca Raton: Taylor & Francis

Group.

Table 1. Data Compilation

Amino acid residue involved and Gbind (Kcal/mol)

morin-3-O--L-
Protein Guaijavarin Quercetin
arabopiranocid

4F55 No interaction on Glu 157 (Gbind-8,8 Glu 157 (Gbind-7,9

protein active site kcal/mol) kcal/mol)


4S3J No interaction on protein binding site

1MPR Arg 8, Arg 33 and Glu Arg 8, Arg 33, Glu 43 Arg 8 and Arg 33

43 (Gbind-9,6 and Asp 116 (Gbind- (Gbind-9,2 kcal/mol)

kcal/mol) 9,0 kcal/mol)

4PH8 No interaction on protein binding site

4L1G Pro 171 (Gbind-6,5 No interaction on Asp 80 (Gbind-7,2

kcal/mol) protein active site kcal/mol)

2HUC Asp 122, Asn 146 and Glu 146, Asn 147, Tyr Glu 146 and His 142

Trp 1 (Gbind-9,2 52 and Ala 3 (Gbind- (Gbind-8,8 kcal/mol)

kcal/mol) 8,2 kcal/mol)

2QFC Asn 201 (Gbind-6,8 Lys 204 (Gbind-6,9 No interaction on

kcal/mol) kcal/mol) protein binding site

4ZVI Asp 73 (Gbind-7,9 Asp 73 (Gbind-7,0 Asp 73 (Gbind-8,0

kcal/mol) kcal/mol) kcal/mol)

4BJP Asn 361, Ser 307, Ser Asn 361, Lys 310, Ser Ser 307, Thr 497 and

359 and Thr 497 307 and Ser 359 Asn 361 (Gbind-7,0

(Gbind-7,3 kcal/mol) (Gbind-7,5 kcal/mol) kcal/mol)

Figure 1. 2QFC (ligan leu-pro-phe-glu-phe)


Figure 2. 4ZVI (ligan 4S4)

Figure 3. Guaijavarin interaction on maltoporin

Figure 4. Quersetin interaction on maltoporin


Figure 5. Guaijavarin interaction on phosphatidylcholine-phospholipase C Bc