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IJPRD, 2011; Vol 5(11): January-2014 (014 - 020)
International Standard Serial Number 0974 – 9446
-------------------------------------------------------------------------------------------------------------------------------------------------DESIGN AND DISCOVERY OF NOVEL THERAPEUTIC NATURAL REPELLENT AGAINST CULEX
QUINQUEFASCIATUS FROM THE PLANT THEVETIA PERUVIANA BY IN SILICO APPROACH
Dhivya R*1 and Manimegalai K
1
Department of Zoology, Avinashilingam Institute for Home Science and Higher Education for Women,
Coimbatore- 641043, Tamil Nadu, India
ABSTRACT
Natural products of plant origin is being tried in the recent past for
control of variety of insect pests and vectors due to the insecticidal,
larvicidal and mosquito repellent properties. Various bioactive
compounds are reported to be present in the plants and
computational techniques play a crucial role in designing and
development of drug of interest. The main function of Odorant
Binding Protein (OBP) is transporting chemical signals
(Semiochemicals) which are identified to be involved in differential
attraction of the insects to the hosts. In the present study potential
ability of the important bioactive compounds from the plant
Thevetia peruviana viz., Peruvianoside I and Thevetin A to block the
odorant receptor proteins of mosquito Culex quinquefasciatus (PDB
id 2L2C) which are well known to recognize semiochemicals from
host is tested and their key role in recognition of host seeking
behaviour was analysed through molecular docking and molecular
dynamics approaches. The present work could provide more
penetrating understanding of the protein- ligand interaction
mechanism. In the present investigation, selected ligands were
docked using Schrodinger Mastero software. The resulys revealed
that Peruvianoside I showed good docking scores with mosquito
odorant binding protein 2L2C and molecular dynamics confirmed
the stability of the bonding. Therefore, it may be used as ideal
candidate for development of potential natural mosquito repellent.
Correspondence to Author
Dhivya R
Department of Zoology,
Avinashilingam Institute for Home
Science and Higher Education for
Women, Coimbatore- 641043, Tamil
Nadu, India
Email: [email protected]
Key words: Molecular docking, Molecular dynamics, Thevetia
peruviana, Culex quinquefasciatus, Ligand, OBP
INTRODUCTION
Mosquitoes are the most important single group of
insects in terms of public health importance, which
transmit a number of diseases, such as malaria,
filariasis, dengue, Japanese encephalitis, etc.
causing millions of deaths every year [1]. The quest
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International Journal of Pharmaceutical Research & Development
to make humans less attractive to mosquitoes has
fuelled decades of scientific research on mosquito
behaviour and control worldwide. Yet mosquitoes
will transmit disease to more than 70 crore people
annually and will be responsible for the deaths of 1
of every 17 people currently alive [2]. Mosquito
borne diseases results from infection with a
protozoan carried by mosquitoes and according to
reports from the World Health Organization
(WHO), it causes as many as 30 lakh deaths
annually[3]. Mosquitoes transmit the arboviruses
responsible for yellow fever, dengue haemorrhagic
fever, epidemic polyarthritis, and several forms of
encephalitis [4].
To prevent proliferation of mosquito borne
diseases and to improve quality of environment
and public health, mosquito control is essential.
The major tool in mosquito control operation is the
application of synthetic insecticides such as
organochlorine and organophosphate compounds.
But this has not been very successful due to
human, technical, operational, ecological, and
economic factors. In recent years, use of many of
the former synthetic insecticides in mosquito
control programme has been limited. It is due to
lack of novel insecticides, high cost of synthetic
insecticides,
concern
for
environmental
sustainability, harmful effect on human health,
other non-target populations, their non
biodegradable nature, higher rate of biological
magnification through ecosystem and increasing
insecticide resistance on a global scale [5,6]. Thus,
the Environmental Protection Act in 1969 has
framed a number of rules and regulations to check
the application of chemical control agents in nature
[7]
. These factors have resulted in an urge to look
for
environment
friendly,
cost-effective,
biodegradable and target specific insecticides
against mosquito species. Considering these, the
application of eco-friendly alternatives such as
biological control of vectors has become the
central focus of the control programmme in lieu of
the chemical insecticides [8].
Relationship between plants and drugs derived
from plants describes the history of mankind.
Plants are important source of natural drugs.
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ISSN: 0974 – 9446
Traditional system of medicine has become a
burning issue of global importance [9]. The
widespread use of synthetic insecticides has led to
many negative consequences resulting in
increasing attention being given to natural
products [10,11]. In modern medicines, plants occupy
a significant place as raw material for some
important drugs, although synthetic drugs and
biotechnology have brought about a revolution in
controlling different disease [12]. Also there is a
growing tendency all over the world, to shift from
synthetic to natural based products including
medicinal and aromatic plants. Less than 5% of the
plants have so far been analyzed as potential
medicine and still there is a great scope of research
in this field [13]. Docking is a term used for
computational schemes that attempt to find the
best matching between two molecules: a receptor
and ligand. The most important concept in drug
design is to understand the methods by which the
active site of a receptor selectively restricts the
binding in appropriate structures [14].
In silico studies help to understand the importance
of small molecules from various plant sources and
their use to enhance protein-ligand interaction.
This approach to screen compounds from plants
depends on various parameters such as size and
shape of the compound and pharmacophoric
groups attached on the compounds, among others.
To assess the efficient therapeutic properties of
novel active compounds from plant origin with
minimum side effects, application of advanced
method like computational techniques play a
crucial role in designing and development of drug
of interest [15]. The present study has been
conducted to analyse the potential ability of the
bio active compounds from the plant Thevetia
peruviana to block the odorant binding proteins of
mosquito Culex quinqufasciatus and also to
understand their key role in inhibiting the host
seeking behaviour of the mosquitoes through
molecular docking or in silico approaches.
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International Journal of Pharmaceutical Research & Development
MATERIALS AND METHODS
Selection of ligands from Calotropis gigantea
Important
potential
bioactive
compounds
[16]
[17]
Peruvianoside I
and Thevetin A
of the plant
Thevetia peruviana selected with the help of
previously published literatures were used in the
present investigation for the computational
prediction of potential drugs from it by the process
of in silico molecular docking.
Molecular docking studies
Target protein retrieval and preparation
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Three dimensional NMR structure of mosquito
odorant binding protein (PDB id: 2L2C) was
obtained from Protein Data Bank (PDB)
(
(Fig 1). The
preparation of a protein involves importing of the
mosquito odorant binding protein structure. The
water molecules have been deleted but water that
bridge between the ligand and the protein were
retained, charges were stabilized, missing residues
were filled in and side chains were generated
according to the parameters available.
Fig 1:: Three dimensional structure of mosquito odorant binding protein (PDB id 2L2C)
Grid generation
Molecular docking of target protein with ligands
Glide was used for receptor grid generation. The
In order to explore the binding mechanism of
prepared mosquito odorant binding protein was
phytochemicals with the target proteins, molecular
displayed in the Workspace. The volume of grid
docking studies have been performed. The two
was calculated. The entire complex was shown
ligands were docked against mosquito odorant
with several types of markers. The enclosing box
binding protein
rotein (2L2C). When the ligand binds with
was made small so that it will be consistent with
protein, the conformation
mation of the protein structure
the shape and character of the protein’s active site
will change and therefore the function of the
and with the ligands that were expected to be
protein will alter automatically. The entire docked
docked.
complex was visualized by using XP visualizer. The
Ligands retrieval and preparation
hydrogen bonding interaction between the
Ligand molecules were retrieved from PubChem
receptor and the ligands were also visualized.
database. The following compounds were retrieved
Molecular dynamic simulation of docked complex:
in 3D SDF format (PubChem id: CID_42608013
42608013 and
In order to confirm the docking results, Molecular
CID_441873).
The two compounds were
Dynamics simulation study was carried out.
processed, unwanted structures were eliminated
Molecular Dynamics simulation was done using
and optimized using LigPrep module from
MacroModel. It is a general purpose, force-ﬁeldforce
Schrodinger.
based molecular modeling program with
applicability to a wide range of chemical systems.
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International Journal of Pharmaceutical Research & Development
MacroModel provides researchers with multiple
advanced methods to understand the chemical
structures, energetics, and dynamics.
Best docked complex was carried for Mo
Molecular
Dynamics. Dynamics is performed using following
parameter such as keeping the constant
temperature at 300 K and in the integration step at
1.0 ps. MD simulations for complex structure was
run. The entire coordinate file was saved every 0 ps
up to 100
00 ps and the result was analyzed by Scatter
Plot.
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Molecular Docking
The 3D structure of odorant binding protein of C.
quinquefasciatus (2L2C) was collected from PDB
PD
(Fig 1). The 3D SDF structures of the two secondary
metabolites viz., Peruvianoside I and Thevetin A of
the plant T. peruviana were taken from PubChem
database. The processed two bioactive compounds
were prepared to dock with the mosquito odorant
binding protein (PDB id 2L2C). The chemical
structures of the two important ligands were
shown in Fig 2
RESULTS AND DISCUSSION
Peruvianoside I
Thevetin A
Fig 2:: 2D structure of ligand compounds retrieved from PubChem database
Among the two ligands docked, the ligand Thevetin
compounds are exhibited in Table 1. Molecular
A didn’t docked with the odorant binding protein.
docking results based on the G-score,
G
H-Bond and
Whereas, the compound Peruvianoside I bound
residue interaction shows binding affinity of the
with the mosquito OBP (PDB id 2L2C). The glide
ligands towards protein 2L2C. If Glide score is
score, number of H-bonds,
bonds, distance of H
H-bonds,
more, the binding affinity of the ligand is higher.
interacted residues and ligand atom of docked
Table 1: Docking Score and H-bond
bond interaction of ligands against mosquito Odorant Binding Protein (PDB id
2L2C)
Sl.
Name of
Compound
G
No. of
Ligand
Distance
Protein residues
No
compound
id
score H bonds
atom
H
GLU 61:(O) O
2.031
1
Peruvianoside I
42608013
-7.55
2
1.922
ALA 62:(O) O
H
2
Thevetin A
441873
The interaction of the compounds with the protein
Peruvianoside I which may have predominant
2L2C at the active site region was confirmed using
repellent ability. Amongst the two compounds in
Schrodinger Mastero software. The results clearly
the present docking study the compound
established high binding affinity of the
Peruvianoside I (Compound id 42608013) produced
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International Journal of Pharmaceutical Research & Development
a good glide score of -7.55 with two Hydrogen
bonds as depicted in Table 4. The two protein
residues were GLU
U 61: (O) O and ALA 62: (O) O
O. The
distance of Hydrogen bonds were 2.031 and 1.922.
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The diagrammatic representation of the
Peruvianoside I docked against mosquito odorant
binding protein (PDB id 2L2C) is shown in Fig 3.
Fig 3: Compound Peruvianoside I docked against mosquito odorant binding protein (PDB id 2L2C)
2L2C
In accordance with the present investigation
performed during 100 ps. The final trajectory files
similar study of in silico docking analysis was
were taken for calculating the Root Mean Square
carried out by Gaddaguti et al [18] for molecular
Deviation of the complex structures.
characterization, phytochemical and docking
While running MD simulation for 42608013
studies of mosquito repellent compounds in
(Peruvianoside I)-2L2C
2L2C complex for 100 ps, the
Ocimum basilicum and showed that N-Hexa
RMSD plot shows the stability of the complex
co
decanoic acid produced good docking scores with
structures at 80ps (Fig 4a).
4
Graphical
mosquito odorant binding protein 3N7H
3N7H.
representation of Time vs. Potential energy map
Compound Thevetin A didn’t bound with mosquito
for 42608013 (Peruvianoside
Peruvianoside I) - 2L2C complex
odorant binding protein. As the compound
structure during molecular dynamics
dynamic simulation for
Peruvianoside I produced an excellent glide score it
100ps is displayed in figure 4b.
4 Tambunan et al [19]
was carried forward for further molecular dynamics
carried out molecular dynamics simulation of DENV
simulation studies.
RNA-dependent RNA-polymerase
polymerase with potential
Molecular Dynamics
inhibitor of disulfide cyclic peptide. The simulation
In order to check the docking results, the Molecular
observation was done by examining the enzymeenzyme
Dynamics simulation was carried out for the
ligand complex interaction between ligand atom
complex of compound 42608013 (Peruvianoside
Peruvianoside I)
with enzyme atom and according to the result of
with the mosquito odorant binding protein (PDB id
simulation; CDEEC is proposed as a better inhibitor
2L2C). Molecular Dynamic (MD) simulation was
of RdRp dengue virus and feasible to be developed
performed to estimate
mate more reliable condition of
as anti-dengue drug.
protein-ligand complex. M D simulation was
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International Journal of Pharmaceutical Research & Development
ISSN: 0974 – 9446
Fig 4: Graphical representation of molecular dynamics simulation studies for docked complex
b)Time vs. Potential energy map for compound
a)Time vs. RMS map for compound 42608013
42608013 (Peruvianoside I) - 2L2C complex
(Peruvianoside I) - 2L2C complex structure for
structure
for 100ps
100ps
The results of molecular dynamics showed that the
studies demonstrated the potiality od the
bonding between the target protein and the
compound Peruvianoside I.. The correlation of our
ligands were stable and got perfectly anchored into
results illustrates the ability of the compound
the active site of the protein. Molecular dynamics
Peruvianoside I in inhibiting the transport of
confirmed the results of molecular docking. When
chemical signals thereby reducing the host seeking
the ligand binds with protein, the conformation of
behaviour of the vectors.
the protein structure will change and therefore the
function of the protein will alter automatically.
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*****
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