In Silico Prediction of High Potential Jararhagin Inhibitor:

Comparison of Batimastat, EDTA and Hydroxytyrosol

Coni A. Kurniasari

, Bayu D. Prakoso

, Eka D. P. Lestari

1,*

and Nia Kurniawan

Biology Department, Faculty of Mathematic and Natural Science,

University of Brawijaya, Malang, 65145, Indonesia

Keywords: Batimastat, EDTA, envenomation, hydroxytyrosol, inhibitor, jararhagin

Abstract: One example of a group P-III SVMP is jararhagin which originates from a Bothrops jararaca. This study

was conducted to compare the possibility of inhibitors that have the highest effectiveness and exact time of

each compound to inhibit hemorragic effect of SVMP. Inhibition of hemorrhagic activity can be done with

several types of compounds that have been known to be inhibitors for SVMP especially jararhagin (PDB

ID: 1C9G) to bind with integrin 

21 (PDB ID: 1AOX). There are batimastat (PubChem ID: 5362422) as

one of peptidomimetic compounds, EDTA (PubChem ID: 6049) as one of zinc chelating agents, and plant

compounds such as hydroxytyrosol (PubChem ID: 82755). The batimastat inhibitory properties from value

of binding energy, found that these inhibitor were more easily bound to jararhagin (-289.0 kcal/mol)

compared to integrin 

21 (-277.1 kcal/mol). That inhibitor also more effectively inhibited by bounding to

jararhagin spread in blood vessels after snakebite because of it’s position and more positive binding energy

(-784.1 kcal/mol). However, unfavorable bonds are formed in the interaction between batimastat inhibitors,

jararhagin and integrin 

21. In inhibitor EDTA interaction, it was found that this compound also more

easily bound to jararhagin (-227.23 kcal/mol), but this inhibitor are more effectively inhibited by bounding

to integrin 

21 because of it’s position and more positive binding energy (-721.57 kcal/mol). In other side

it also has unfavorable bonds. While the interaction of hydroxytyrosol shows that inhibitor are easier to

interact with jararhagin and more effectively acts as a jararhagin inhibitor by being consumed after the body

is exposed to jararhagin (-781.33 kcal/mol) without showing an unfavorable bond. We can conclude that the

natural inhibitors formed in hydroxytyrosol from olive oil are more stable and have highest possibility in

preventing hemorrhagic symptoms due to snake bites that contain jararhagin venom.

1 INTRODUCTION

Envenomation is one of dangerous health problem

because of the death risk. There are 5,5 millions

envenomation cases annually. Snake venom contains

mixture of various proteins or protein families with

different bioactivities and tissue target (Williams

et.al., 2010). The examples of protein families in

snake venom are snake venom metalloproteinase

(SVMP), snake venom serine proteinase (SVSP),

cysteine-rich secretory protein (CRiSP),

phospholipase A

2, phospholipase type B, C-type

lectin-like protein, L-amino acid oxidase (LAAO), 3

finger toxin (3FTx), et cetera (Kunalan et.al., 2018).

SVMP (Snake Venom Metalloproteinase) is one

of protein family in Elapidae and Viperidae snake

venom. Up to 30% Viperidae venom is consist of

SVMP (Silva et.al., 2016).

SVMP is a zinc-dependent hydrolase which has

catalytic zinc ion in the active site. The catalysis

process of this enzyme needs zinc ion (Zn

) as the

mediator, zinc ion is coordinated with 3 side chain

of histidine and water molecule binded with

glutamate residue (Preciado et.al., 2018). SVMP has

the hemorrhagic effect and fibrynogenolytic. It can

cleave A and B chain on fibrynogen. SVMP can

degrade some of extracellular matrix proteins i.e.

collagen IV, laminine, fibronectin, and proteoglycan

perlecan. SVMP can act as the mediator of local

tissue damage, and induce the endothelial cell

hemorrhage as well. The damage of endothelial cell

and basal membrane on blood vessels will helps the

toxic protein spread to the tissue target

(Pithayanukul et.al., 2009). Jararhagin is a member

of SVMP protein family with high hemorrhagic

effect. It is a 52 kDa PIIIb SVMP, the first

Kurniasari, C., Prakoso, B., Lestari, E. and Kurniawan, N.

In Silico Prediction of High Potential Jararhagin Inhibitor: Comparison of Batimastat, EDTA and Hydroxytyrosol.

DOI: 10.5220/0009586800050014

In Proceedings of the 6th International Conference on Advanced Molecular Bioscience and Biomedical Engineering (ICAMBBE 2019) - Bio-Prospecting Natural Biological Compounds for

Seeds Vaccine and Drug Discovery, pages 5-14

ISBN: 978-989-758-483-1

metalloproteinase isolated from B. jararaca

(Ferreira et.al., 2018).

The specific treatment of envenomation case is

conducted by antivenom treatment. According to

WHO (2010), antivenom or antivenin is consisted of

pure immunoglobulin fragment from animal plasma

which have been immunized by snake venom.

Antivenom treatment has several disadvantages.

Antivenom only neutralize any of the venoms used

in its production, or from closely related species. It

needs suitable storage condition because of the

sensitive components in antivenom. It is unsuitable

to neutralize the local tissue damage (Preciado et.al.,

2018). Antivenom has high effectivity in

neutralizing systemic effect of a venom, but it has

low effectivity in neutralizing local effect of snake

venom, i.e. effect of SVMP (Romero et.al., 2012).

Enzyme inhibitor has become a potention to treat

local tissue damage effect of SVMP. Some

compounds which are known as inhibitor of SVMP

are peptidomimetic, zinc chelating agent, and

phenolic compound. Peptidomimetic such as

Batimastat and Marimastat; zinc chelating agents

such as EDTA, DTPA, TTD; and phenolic

compounds are known as SVMP enzyme inhibitor

(Preciado et.al., 2018). But there is no knowledge

about which compound is most effective to inhibit

SVMP hemorrhagic activity. The aim of this study is

to compare Batimastat (peptidomimetic), EDTA

(zinc chelating agent), and hydroxytyrosol (phenolic

compound) as the most effective inhibitor of

jararhagin SVMP.

2 MATERIALS AND METHODS

2.1 Protein and Ligand Structure

Preparation

We use the structure of 2 proteins, the snake venom

metalloproteinase (SVMP) jararhagin (PDB ID:

1C9G) and α

2β1 intregin (PDB ID: 1AOX). Protein

structure was downloaded from RCSB PDB

database (https://www.rcsb.org/). We also use

structure of 3 ligands, Bastimastat (PubChem ID:

5362422), EDTA (PubChem ID: 6049), and

Hydroxytyrosol (PubChem ID: 82755). Ligand

structure was downloaded from PubChem database

(https://pubchem.ncbi.nlm.nih.gov/). Preparation of

protein structure was conducted using Discovery

studio 16.1.0 software. Each protein (jararhagin and

2β1 intregin) was prepared by deleting the water

and ligand molecule. Protein structures then saved as

PDB format (.pdb). The ligand structure was

prepared using PyRx software. Ligands was

prepared by minimizing free energy and converted

to PDB format.

2.2 Docking and Visualization

The docking of jararhagin and ligand with α2β1

integrin collagen receptor on the cell-surface of

platelet were carried out in two types of conditions

to determine the effectiveness and efficiency of

inhibitors. The first condition is interaction between

jararhagin-ligand complex and α

2β1 integrin. The

second condition is interaction between α

2β1

integrin-ligand complex and jararhagin. Since we

used 3 kind of ligands (Batimastat, EDTA, and

hydroxytyrosol), there are 6 types of interaction

between jararhagin, ligand, and α

2β1 integrin. The

docking was performed using HEX 8.0.0. Docking

results were visualized using Discovery studio

16.1.0.

3 RESULT AND DISCUSSION

3.1 The Interaction of Jarharagin with

Batimastat and Α2β1 Integrin

Docking energy between jararhagin (green) and

batimastat (red) ligand is -289.0 kcal/mol. Whereas

if the results of jararhagin-batimastat docking are re-

docked with the integrin receptor α

2β1 (α: blue, β:

violet) , the docking energy decreases with a value

of -784.1 kcal/mol. The interaction between

(jararaghin + batimastat) and integrin α

2β1 are a first

condition that indicated treatment of batimastat after

envenomation. This condition has 10 of favorable

bonds. Two conventional hydrogen bonds that

binding the amino acid residues of Leu53 with atom

H and O on Ligan 1. Then 2 carbon-hydrogen bonds

between Ligand with the amino acid residues Leu53

and Tyr53. One Pi bond with sulfur with amino acid

residue Tyr7. One Pi bond with a lone pair is Asn38.

Three hydrophobic bonds with a Pi type with alkyl

are all three Lig1 with Ala52, Pro202, and Ala37.

One type hydrophobic bond between Pi is amino

acid residue Tyr7. There are also appear 6 pieces

that are unfavorable, which is appear in 2 kind of

amino acid residue like Asn42 and His50.

The interaction between jararhagin and the

docking results of batimastat and integrins α

2β1. The

docking energy of the batimastat and integrin α2β1

is -277.1 kcal/mol. If the result of the docking is

docked again with a fault, the value of the docking

energy becomes -857.5 kcal/mol. The interaction of

ICAMBBE 2019 - 6th ICAMBBE (International Conference on Advance Molecular Bioscience Biomedical Engineering) 2019

jararhagin and (Batimastat + Integrin α2β1) are

second condition that indicate treatment of

batimastat before envenomation. This condition has

4 favorable binding bonds, each of which has a

different type of bond. The first bond is a

conventional hydrogen bond that binds with amino

acid residue Asn27, then carbon bonds with

hydrogen that binds atom H with amino acid

residues Ala245, alkyl bonds that bind atom C with

amino acid Leu276, sulfur bond with X which binds

atom S with Glys338 amino acid residu. The last,

there are also 3 unfavorable bond that appeat in 2

kinds of amino acid residues like Gly338 and

Asn274. The docking results between jararhagin and

integrin wich are a presumed condition if jararhagin

envenomation occure. produce docking energy of -

841.3 kcal/mol.

The result show that batimastat is easier to

interact with jararhagin than the integrin receptor

α2β1 because the energy used to interact with

jararhagin is smaller than the interaction with the

integrin receptor α2β1. Morover, the docking

condition 2 that is batimastat-intgerine complex

α2β1 interaction with jararhagin has a lower

(negative) docking energy value of -841.3 kcal / mol

compared with the jararhagin-batimastate complex

interaction with the α2β1 integrin the value is higher

(positive) which is -784.1 kcal / mol. So that the

batimastatic-integrin α2β1complex that was

interacted with jararahgin was less effective in

inhibiting the enzyme formation of jararhagin

because batimastate was easier to tie jararhagine

than before binding to the integrin α2β1.Lower

docking energy shows that the bonds between

protein requires more energy to bind, so that it can

be used to inhibit jararhagin for integrin receptors.

In addition, when viewed from a 3-dimensional

structure, it can be seen from the 3-dimensional

interaction that the batimastat ligand is positioned

between jararhagin and the integrin domain α2

(Figure 1F). This is in accordance with the reference

which states that jararhagin will bind to integrins in

the α2 domain then integrin β1 cleavage, even with

low docking energy that allows easy unbonding.

Batimastat inhibits jararhagin which contains ZBG

or zinc binding group by cleavage BaP1 protein

between dermal-epidermal formed from basic

membrane components. High docking energy also

shows that the bonds between molecules are strong

so they are not easily released (Jimenez et.al., 2008).

Integrin-binding motif α

2β1 is located to or within

the hyper-variable region of the cycstein-rich

domain. Part that causes inhibition of platelet

aggregation is the catalytic or proteolytic site that

interacts with the integrin α

2β1 so that it will trigger

signal transduction on platelets (Tanjoni et.al.,

2010).

The atomic interactions between batimastats and

jararhagin interact more than the atomic interactions

between batimastats and integrins α2β1. The

interaction of the jararhagin-batimastat complex

with integrins α2β1 has more hydrophobic and

hydrogen bonds than the complex interactions of

jararhagin-integrin α2β1 with jararhagin.

Conventional hydrogen bonds are stabilizing bonds

in biomolecular structures. Hydrogen bonds occur

between proton donor groups. The donor part is an

electronegative element and the acceptor group is a

free electron pair or phi bond, especially on oxygen

and nitrogen atoms (Horowitz & Trievel, 2012).

Figure 1. Interaction of Jararhagin, Batimastat, and

Integrin 

21. A. Ligan interaction between complex

jararhagin-batimastat and integrin 

21 (3D). B. Ligan

interaction between complex jararhagin-batimastat and

integrin 

21 (2D). C. Ligan interaction between complex

integrin 

21 -batimastat and jararhagin (3D). D. Ligan

interaction between complex integrin 

21 -batimastat and

jararhagin (2D). E. Interaction complex jararhagin-

batimastat and integrin 21. F. Interaction complex

integrin 

21-batimastat and jararhagin.

C D

In Silico Prediction of High Potential Jararhagin Inhibitor: Comparison of Batimastat, EDTA and Hydroxytyrosol

Table 1. Bonding interactions between the jararhagin batimastat complex and the integrin receptor α2β1.

Name

Distance

Category

Type

From

(Å)

Chemistry

A:LEU53:HN -

2,64948Å

Hydrogen

Conventional

H-Donor

H-Acceptor

A:LIG1:O Bond

Hydrogen Bond

A:LIG1:H -

2,89807Å

Hydrogen

Conventional

H-Donor

H-Acceptor

A:LEU53:O

Bond

Hydrogen Bond

A:ALA52:HA -

2,37972Å

Hydrogen

Carbon Hydrogen

H-Donor

H-Acceptor

A:LIG1:O Bond

Bond

A:LIG1:H -

2,13003Å

Hydrogen

Carbon Hydrogen

H-Donor

H-Acceptor

A:LEU53:O Bond

Bond

A:LIG1:S -

4,163Å Other

Pi-Sulfur

Sulfur Pi-Orbitals

A:TYR7

A:ASN38:OD1 -

2,82212Å Other Pi-Lone Pair

Lone Pair

Pi-Orbitals

A:LIG1

A:LIG1 -

4,23106Å

Hydrophobic

Pi-Pi Stacked

Pi-Orbitals

A:TYR7

A:LIG1 -

5,49552Å

Hydrophobic

Pi-Alkyl

Pi-Orbitals Alkyl

A:ALA52

A:LIG1 -

4,09041Å

Hydrophobic

Pi-Alkyl

Pi-Orbitals Alkyl

A:PRO202

A:LIG1 -

4,39295Å

Hydrophobic

Pi-Alkyl

Pi-Orbitals Alkyl

A:ALA37

A:ASN41:ND2 -

1,84028

Unfavorable Unfavorable Bump Steric

Steric

A:LIG1:O

A:ASN41:ND2 -

1,46595

Unfavorable Unfavorable Bump Steric

Steric

A:LIG1:H

A:ASN41:HD21

1,76436

Unfavorable Unfavorable Bump Steric

Steric

- A:LIG1:O

A:ASN41:HD22

1,45353

Unfavorable Unfavorable Bump Steric

Steric

- A:LIG1:O

A:ASN41:HD22

Unfavorable

Steric;H- Steric;H-

0,646487

Unfavorable

Bump;Unfavorable

- A:LIG1:H

Donor Donor

Donor-Donor

A:HIS50:HD1 -

1,52363

Unfavorable

H-Donor

A:LIG1:H Donor-Donor

ICAMBBE 2019 - 6th ICAMBBE (International Conference on Advance Molecular Bioscience Biomedical Engineering) 2019

Table 2. Bonding interactions between the batimastat integrin α2β1 complex and jararhagin.

Name

Distance

(Å)

Category TYPE

From

Chemistry

A:ASN274:HD21

- B:LIG1:O

2,1888Å

Hydrogen

Bond

Conventional

Hydrogen Bond

H-Donor

H-

Acceptor

B:LIG1:H -

A:ALA245:O

1,6891Å

Hydrogen

Bond

Carbon

Hydrogen Bond

H-Donor

H-

Acceptor

B:LIG1:S -

A:GLY338:N

2,4186Å Other Sulfur-X Sulfur O,N,S

B:LIG1:C -

A:LEU276

4,5753Å Hydrophobic Alkyl Alkyl Alkyl

A:ASN274:HB1 -

B:LIG1:O

1,67052 Unfavorable

Unfavorable

Bump

Steric Steric

A:GLY338:CA -

B:LIG1:O

2,23482 Unfavorable

Unfavorable

Bump;Carbon

Hydrogen Bond

Steric;H-

Donor

Steric;H-

Acceptor

A:GLY338:HN -

B:LIG1:S

1,83166 Unfavorable

Unfavorable

Bump

Steric Steric

The results of research on hemorrhagic ability by

jararhagin in the lungs and skin with experimental

animals showed that batimastat was able to reduce

hemorrhagic activity in these organs. Other results

showed that jararhagin incubated with batimastat

and inserted into intradermal mice showed that there

was a reduction in hemorrhagic diameter in

experimental animals compared to incubation of

jararhagin with human 2-macroglobulin and normal

serum mouse (Escalante et.al., 2003).

3.2 The Interaction of Jararhagin with

Α2β1 Integrin and EDTA

The first condition is the interaction between

jararhagin-EDTA complex which requires an energy

of -227.23 kcal/mol to bound each other. This

interaction are made by α

2β1 integrin receptor which

is a common target of the inhibition process by

jararhagin. The energy needed from the ligand

complex jararhagin-EDTA to bound with I domains

of α

2 (colored blue) is -733.96 kcal/mol. While the

position of EDTA is area that does not interact with

integrin amino acid residues (jararhagin is green)

(Figure 2E). The second condition is an alternative

interaction between the α

2β1-EDTA complex which

requires higher energy, which is -215.46 kcal/mol to

bound with I domain of α

2. That interaction made

with jararhagin being the natural inhibitor of the

integrin receptor α

2β1. The energy required from the

receptor complex α

2β1-EDTA to bound with the

disintegrin-like domain of jararhagin (green) is -

721.57 kcal/mol (Figure 2F). This condition showed

that EDTA is more easily bound with jararhagin

compared to α

2β1 integrin. In addition, when

jararhagin-EDTA are the ligand complexed it

required lower energy to bound to platelet integrin

surface receptors, compared to the energy needed

when EDTA in the form of receptor complexed with

integrins. While the tendency of bond energy and

bounding-site position between the receptor complex

and jararhagin indirectly indicates that inhibition by

EDTA is more effective by bounding to the integrin

2β1 even before the venom spreads in the blood

vessels after the envenomation.

Furthermore, the interaction between the

jararaghin-EDTA ligand complex and 

21 integrins

forms 5 kind of favorable bonds (Figure 2F). Four of

them are conventional hydrogen bonds with an

average distance of 2.3 Å to 2.9 Å that can be

classify as strong until medium H-bond. Strong

covalent H-bond have length 2.2 Å - 2.5 Å whereas

moderate mostly electrostatic have length of distance

2.5 Å -3.2 Å (Baey, 2013). The hydrogen bond

bound to asparagine amino acid residues (Asn194),

proline (Pro195), tyrosine (Tyr5), and aspartic acid

(Asp3). Then also formed one carbon hydrogen bond

is formed at A LIG: 1. A hydrogen bond interaction

span a large interval, ranging from tiny energies to

large values when the acceptor is an anion that can

devise interaction stability. Hydrogen bond is

generally also stronger interaction, but still less

stable than van der Waals interaction cause hydrogen

bond have shorter distance (Mingos, 2004). A

hydrogen bond can be called conventional or

classical if it is formed between a partly positively

charged hydrogen atom in proton-donor component

and the lone electronic pair of electronegative

element acting as a proton-accepting component.

This conventional hydrogen bonds forming from

weak to medium energy and accompanied by a

remarkable interpenetration (Bakhmutov, 2008). In

In Silico Prediction of High Potential Jararhagin Inhibitor: Comparison of Batimastat, EDTA and Hydroxytyrosol

addition, almost same as the results of interactions

with batimastat inhibitors, the interaction of EDTA

inhibitor ligand complex also forms 4 unfavorable

bonds, that appear namely between ligands in 2 kind

of amino acid residues like proline (Pro4) and lysine

(Lys6). This unfavorable bump interaction bond in

the wrong area that can cause the interaction

unstable. Unfavorable bump are generally formed by

tripled carbon interaction (Karimi & Nalapogaja,

2012).

The interaction of jararhagin with the EDTA-

integrin receptor complex 

21 forms 8 kind of

favorable bonds (Figure 2D). The first type of bond

is a conventional hydrogen bond which consists of 5

bonds, namely ligand bonds with serine amino acid

residues (Ser244), alanine (Ala245), lysine (Lys247),

asparagine as a donor and H receptor ( Asn274). The

five bonds show a distance of 1.7 Å to 3 Å. The

second type of bond is carbon hydrogen bonds with 1

bond between ligands with serine amino acid

residues (Ser138) and 2 bonds between amino acids

alanine (Ala245) with bond distances ranging from

1.7 Å to 3.7 Å that can be classify as strong until

weak H-bond. Strong covalent H-bond have length

2.2 Å - 2.5 Å whereas moderate mostly electrostatic

have length of distance 2.5 Å - 3.2 Å, and the weak

electrostatic dispersed have length of distance more

than 3.2 Å (Escalante et.al., 2003). While the last

type of bond is 5 unfavorable bonds, that appear

namely between ligands in 3 amino acid residues like

alanine (Ala245), arginine (Arg243) and tyrosine

(Try 235). This bond shows that the formed

interaction is less stable even though it successfully

inhibits the bounding of jararhagin to the position of

domain I α

2 according to the form of the disturbance

caused by the ability of jararhagin to block integrin

interactions of 

21 with collagen by bounding to α2

domain I or by cleavage of 

21 [16]. This inhibitor

also proven to be the most effective in vivo and in

vitro method to irreversibly inactivate the proteolytic

activity of jararhagin by remove the active site zinc

and structural calcium molecules from the protein

using EDTA (Gallagher et.al., 2005).

3.3 The Interaction of Jararhagin with

α2β1 Integrin and Hydroxytyrosol

The binding energy of jararhagin-hydroxytyrosol is -

178.3 kcal/mol. The interaction between jararhagin-

hydroxytyrosol and α2β1 integrin needs -781.33

kcal/mol to bind. Jararhagin-hydroxytyrosol is

binding α2β1 integrin on 2-I domain (symbolized

with blue coloration on Figure 3), the inhibitor is not

attached on the interaction site between α2β1

integrin and jararhagin.

On the other hand, the docking in second

condition showed that interaction between

hydroxytyrosol and α2β1 integrin has binding

energy -166.7 kcal/mol. Hydroxytyrosol as ligand

bind at the 2-I domain of α2β1 integrin. The

interaction between α2β1 integrin-hydroxytyrosol

and jararhagin needs -809.3 kcal/mol to bind. The

binding position of hydroxytyrosol is located near to

binding target site of jararhagin generally. The

energy yielded from interaction of α2β1 integrin-

hydroxytyrosol is more than the interaction of

jararhagin-hydroxytyrosol. It is showed that

hydroxytyrosol is easier to bind with jararhagin than

α2β1 integrin after the envenomation.

Figure 2. Interaction of Jararhagin, EDTA, and Integrin



21. A. Ligand interaction between complex jararhagin-

EDTA and integrin 

21 (3D). B. Ligand interaction

between complex jararhagin-EDTA and integrin 

21

(2D). C. Ligand interaction between complex integrin



21-EDTA and jararhagin (3D). D. Ligand interaction

between complex integrin 21-EDTA and jararhagin

(2D). E. Interaction complex jararhagin-EDTA and

integrin 

21. F. Interaction complex integrin 21-EDTA

and jararhagin.

ICAMBBE 2019 - 6th ICAMBBE (International Conference on Advance Molecular Bioscience Biomedical Engineering) 2019

Table 3. Detail information of H-Bond interaction between jararhagin-EDTA ligands complex with α2β1 integrin.

Name

Distance

Category

Types

From Chemistry To Chemistry

(

)

A:LIG1:H - 2,98453 Hydrogen Bond Conventional

H-Donor

H-Accepto

A:ASN194:O

Hydrogen Bond

A:LIG1:H - 2,84653 H

dro

en Bond Conventional

H-Donor

H-Acce

A:PRO195:O

Hydrogen Bond

A:LIG1:H - 2,81501 Hydrogen Bond Conventional

H-Donor

H-Accepto

A:TYR5:O

Hydrogen Bond

A:LIG1:H - 2,57797 H

dro

en Bond Conventional

H-Donor

H-Acce

A:ASP3:OD1

Hydrogen Bond

A:ASP3:CA -

3,2238

dro

en Bond Carbon H

dro

H-Donor

H-Acce

A:LIG1:O

Bond

A:PRO4:CD -

Unfavorable

Steric;H-

2,09393 Unfavorable Bump;Carbon

Steric;H-Donor

A:LIG1:O Acceptor

Hydrogen Bond

A:PRO4:HD1 -

1,53485 Unfavorable

Unfavorable

Steric Steric

A:LIG1:O Bump

A:LYS6:CG -

2,19754 Unfavorable

Unfavorable

Steric Steric

A:LIG1:O Bump

A:LYS6:CD -

2,19833 Unfavorable

Unfavorable

Steric Steric

A:LIG1:O Bump

Table 4. Detail information of H-Bond interaction between α2β1-EDTA receptors complex with jararhagin.

Name

Distance

Category Types

From Chemistry

(

)

Chemistry

A:SER244:HN - 1,85267 Hydrogen Conventional

H-Donor

H-Accepto

B:LIG1:O

Bond

Hydrogen Bond

A:ALA245:HN -

2,62916 H

dro

en Conventional

H-Donor

H-Acce

B:LIG1:O

Bond

Hydrogen Bond

A:LYS247:HN -

1,76284 Hydrogen Conventional

H-Donor

H-Accepto

B:LIG1:O

Bond

Hydrogen Bond

A:ASN274:HD21

3,033 H

dro

en Conventional

H-Donor

H-Acce

- B:LIG1:O

Bond

Hydrogen Bond

B:LIG1:H -

2,45232 H

dro

en Conventional

H-Donor

H-Acce

A:ASN274:O

Bond

Hydrogen Bond

B:SER138:CB -

3,76774 H

dro

Carbon H

dro

H-Donor

H-Acce

B:LIG1:O

Bond Bond

B:LIG1:H -

2,24188 Hydrogen

Carbon Hydrogen

H-Donor

H-Accepto

A:ALA245:O

Bond Bond

B:LIG1:H -

1,73094 Hydrogen

Carbon Hydrogen

H-Donor

H-Accepto

A:ALA245:O

Bond Bond

A:TYR235:O -

1,93483 Unfavorable Unfavorable Bump

Steric Steric

B:LIG1:O

A:ARG243:N -

1,99267 Unfavorable Unfavorable Bump

Steric Steric

B:LIG1:O

A:ARG243:HN -

Unfavorable

Steric;H-

1,6825 Unfavorable

Bump;Conventional Steric;H-Donor

B:LIG1:O

Acceptor

Hydrogen Bond

A:ALA245:O -

2,07184 Unfavorable Unfavorable Bump

Steric Steric

B:LIG1:C

B:LIG1:H -

Unfavorable

Steric;H-

1,27798 Unfavorable Bump;Carbon

Steric;H-Donor

A:ALA245:O

Acceptor

Hydrogen Bond

In Silico Prediction of High Potential Jararhagin Inhibitor: Comparison of Batimastat, EDTA and Hydroxytyrosol

When α2β1integrin-hydroxytyrosol interacted to

jararhagin, the binding energy is less than energy of

interaction between jararhagin-hydroxytyrosol and

α2β1 integrin. It is showed that hydroxytyrosol as

the inhibitor would inhibit effectively if it is

consumed after envenomation.

Hydroxytyrosol is one of phenolic compound

which has the high level of antioxidant (Obied et.al.,

2012). Interaction of phenolic compound and SVMP

will form the hydrogen bond with three histidine

residue on zinc binding motive area. Thus, zinc ion

will be chelated from SVMP complex. Zinc ion is an

important component of SVMP, because SVMP is

categorized as zinc-dependent hydrolase. When zinc

ion is chelated from SVMP, its enzimatic activity is

inhibited (Pithayanukul et.al., 2009).

Interaction of jararhagin-hydroxytyrosol and



21 integrin is consist of 1 hydrogen bond and 2

hydrophobic bonds. The first bond is hydrogen bond

which formed from hydrogen atom on ligand to

Pro202 residue as hidrogen receptor. The distance of

this bond is 2,88431. The second bond is

hydrophobic bond (Pi-Pi Stacked) which formed

from Tyr7 residue of jararhagin protein (to an atom

of ligand. The distance of this bond is 4,14095. The

third bond has same type as the second bond, which

formed from His50 residue to an atom of

hydroxytyrosol. The distance of this bond is

4,82085.

Pi-pi (-) stack is a type of non-covalent bond.

That type of bond is formed between two aromatic

ring from different compound. It has acquainted for

its role to stabilize the macromolecular structures

such as nucleic acid, protein, and other material

(Boehr et.al., 2002). Thus, the presence of two Pi-Pi

stacked hydrophobic bond, indicate the strong

interaction between jararhagin-hydroxytyrosol and



21 integrin.

On the other hand, interaction between 

21

integrin-hydroxityrosol with jararhagin has only one

hydrogen bond. This bond is formed from hydrogen

atom on ligand with Asn274 residue. The distance of

this bond is 1,90548. Hydrogen bond is the

interaction between hydrogen atom and

electronegative atom group, it has stronger bond

than van der Waals interaction, and weaker than

covalent or ionic bond. Hydrogen bond cosidered to

be the regulator of protein-ligand binding. This bond

can create stronger protein-ligand interaction but

causing absence of net gain binding affinity, but this

bond is also reported to enhance ligand binding

affinity by displacing protein-bound water molecule

to the bulk solvent (Chen et.al., 2016).

Since there is no any unfavorable bond, the

interaction with phenolic compound is better than

other ligand because of the stability. Besides, this

natural inhibitor is more efficient because it include

the metal chelator activity, high level of antioxidant

which can support the cell regeneration, free radical

scavenger, enzyme activity modulator, and

anticancer (Pithayanukul et.al., 2009).

Figure 3. Interaction of Jararhagin, Hydroxytyrosol, and

Integrin 

21. A. Ligan interaction between complex

jararhagin-hydroxytyrosol and integrin 21 (3D). B. Ligan

interaction between complex jararhagin-hydroxytyrosol

and integrin 

21 (2D). C. Ligan interaction between

complex integrin 

21 - hydroxytyrosol and jararhagin

(3D). D. Ligan interaction between complex integrin 

21

-hydroxytyrosol and jararhagin (2D). E. Interaction

complex jararhagin-hydroxytyrosol and integrin 

21. F.

Interaction complex integrin 

21-hydroxytyrosol and

jararhagin.

ICAMBBE 2019 - 6th ICAMBBE (International Conference on Advance Molecular Bioscience Biomedical Engineering) 2019

Table 5. Interaction between jararhagin, hydroxytyrosol, and 21 integrin.

Name

Distance

Category Types

From

(Å)

Chemistry Chemistry

Interaction

A:LIG1:H -

Hydrogen

Conventional

between

2,88433 Hydrogen

H-Donor H-Acceptor

A:PRO202:OXT

Bond

jararhagin-

Bond

hydroxytyrosol

ligands

A:TYR7 -

4,1407

Hydrophobic Pi-Pi Stacked Pi-Orbitals Pi-Orbitals

complex with

A:LIG1

21 integrin

A:HIS50 -

4,8207

Hydrophobic Pi-Pi Stacked Pi-Orbitals Pi-Orbitals

A:LIG1

Interaction

between 21

integrin -

B:LIG1:H -

Hydrogen

Carbon

hydroxytyrosol

1,90548 Hydrogen

H-Donor H-Acceptor

A:ASN274:O

Bond

ligands

Bond

complex with

jararhagin

4 CONCLUSIONS

Inhibition of hemorrhagic activity can be done with

several types of compounds that have been known to

be inhibitors for SVMP especially jararhagin can

done by batimastat (peptidomimetic compounds),

EDTA (zinc-chelating agents), and plant compounds

such as hydroxytyrosol. Based on the results of the

in silico analysis from batimastat, EDTA and

hydroxytyrosol inhibitory properties, found that

these inhibitor were more easily bound to jararhagin

compared to integrin 

21. But only two of them are

more effectively inhibited

by bounding to jararhagin

spread in blood vessels after snakebite cases.

However, unfavorable bonds are formed during

interaction between batimastat inhibitors, jararhagin

and integrin 

21. These inhibitor are Batimastat and

hydroxytyrosol. Furthermore, in the second inhibitor

EDTA, it was found that this compound more

effective doing inhibition by inhibiting integrin 

21.

In other side inhibitor batimastat and EDTA also

has

unfavorable bonds. While the last alternative of

phenolic compounds in the form of hydroxytyrosol

shows that inhibitor are interact with jararhagin and

integrin α

2β1 without showing an unfavorable bond.

From that result we can conclude that the natural

inhibitors formed in hydroxytyrosol from olive oil

are more stable and have highest effectiveness and

efficiency in preventing hemorrhagic symptoms due

to snake bites that contain jararhagin venom.

ACKNOWLEDGEMENTS

The present study was funded by Kemenristekdikti -

the Government of the Republic of Indonesia

through the scheme of PDUPT 2019 to Nia

Kurniawan with contract number

330.13/UN10.C10/PN/2019.

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ICAMBBE 2019 - 6th ICAMBBE (International Conference on Advance Molecular Bioscience Biomedical Engineering) 2019