INTRODUCTION:
The
mitogen-activated protein kinase (MAPK) pathway is a vibrant channel
transmitting and amplifying extracellular signals to intracellular components
and play a pivotal role in cell proliferation, growth and survival process.
Growth factors interact with their specific receptors and activate receptor
tyrosine kinase (RTK) family, thus amplifying and transducing signal cascade
through cytosolic and nuclear intermediates, to regulate gene expression1,2,3.
Cyanidin belongs to anthocyanins, a
flavonoid that imparts brilliant colour to fruits and flowers4,5.
Based on the previous studies it is
established that it has antioxidant activity5-16, anti-inflammatory
activity6,12,13,16-20, anti-apoptosis activity6,21-26,
anti-mutagenic activity4,13,27 and anti-carcinogenic activity4,12-15,28.
Being one of the major groups of naturally occurring anthocyanins with good
anticancer, antioxidant and free radical scavenging activity the mode of these
action of cyanidin is poorly established. Hence we propose that cyanidin may
exhibit these activity by modulating the MAPK pathway4. Thus the aim
of our present study was to determine the effect of cyanidin on molecular
proteins of MAPK pathway by insilico docking using Auto dock 4.2.
MATERIALS AND
METHODS:
Preparation of cyanidin:
The structure of cyanidin (Pubchem Id: 128861) was
imported from PubChem in sdf format and drawn in Marvin sketch. The sdf file
was converted to pdb format using Open Babel-2.3.1 and finally it was saved in
pdb format.
Preparation of MAPK pathway proteins:
The MAPK pathway
proteins such as epidermal growth factor (EGF) (1EDM), fibroblast growth factors (FGF) (1RG8), platelet-derived growth factor receptor (PDGFR)
(1H9O), receptor tyrosine kinase (RTK) (2Y1N),
RAS (4LPK), rapidly accelerated fibro sarcoma
((RAF) (4H29), mitogen activated protein kinase kinase (MEK) (6YFZ),
extracellular signal-regulated kinases (ERK) (3SA0), jun proto-oncogene,
AP-1 transcription factor subunit (JUN) (4H39), fos proto-oncogene, AP-1 transcription factor subunit (FOS)
(1FXL), Son of Seven less (SOS) (2II0) and mitogen-activated protein kinase
(MAPK) (3S4E) were imported from RCSB Protein Data Bank in PDB format with
their respective PDB ID. Cyanidin was docked with the above 12 molecular
proteins of MAPK pathway using Auto Dock Tools 4.2 (Version 1.5.6). The
heteroatoms of the proteins were removed and was saved in the pdb format. The
interaction activity of cyanidin against MAPK molecular proteins was studied
using Auto dock 4.2.
Molecular Docking studies by Auto Dock Tools 4.2
(Version 1.5.6):
The receptor and cyanidin interactions was predicted
using Auto Dock 4.2 with certain modification. The proteins were prepared by
adding all hydrogen atoms (polar only) to the macromolecule to correct the
calculation of partial atomic charges. Gasteiger charges were calculated for
each of the atom in the MAPK molecule. 3-D affinity grids of size 60*60*60 Å
with 0.375 Å spacing were centered on the geometric center of the target protein.
The dimensional affinity grid was fixed as specified above, to run the Auto
grid file. The selected important docking parameters for the Lamarckian Genetic
Algorithm (LGA) are as follows: population size of 150 individuals, mutation
rate of 0.02, crossing over of 0.8, 25 docking runs, random initial positions
and conformations29,30,31,32. The cyanidin was docked with MAPK
pathway proteins using Auto Dock 4.2 (Version 1.5.6) software with the above
mentioned procedure. The binding energy, inhibition constant, number of
hydrogen bonds and active residues interaction were simulated.
Out of 12 molecular proteins of the MAPK pathway, 11
molecules (Table 1) namely EGF, FGF, PDGF, RTK, RAS, MEK, RAF, ERK, JUN, FOS
and SOS exhibited favourable binding energy above (-5kcal/mol) and formed
nearly 1-3 hydrogen bonds. MAPK did not exhibit favourable binding energy.
Cyanidin exhibited (Figure 1) good inhibition constant of 215.32
m with 1 hydrogen bond and binding energy
of -5.00kcal/mol for PDGFR. Cyanidin also exhibited inhibition constant of
above 11.64µm and 13.48µm for RAF and ERK respectively. Cyanidin did not show
favourable interaction with MAPK.
Figure.1 Cyanidin docked with PDGFR
Figure.2 Cyanidin docked with ERK
Figure.3 Cyanidin docked with RAF
DISCUSSION:
Our study showed
that cyanidin acts through MAPK pathway by modulating EGF, FGF, PDGF, RTK, RAS,
MEK, RAF, ERK, JUN, FOS and SOS (Table 1) proteins. Cyanidin are flavonoids
with oxonium ion (flavylium) and studies report that their biological
activities are due to their free radical scavenging and antioxidant activity.
The catechol structure of cyanidin shows high antioxidant activity due to the
formation of stable semi quinone radicals which is characterized by electron
and hydrogen atom donating activity33. The resulting radicals can
also be stabilized by hydrogen bonding with neighbouring hydroxyl group34.
Thus the antioxidant and free radical scavenging activity of cyanidin may contribute
to this activity.
Cyanidin exhibited (Figure 1) good inhibition constant
of 215.32µm with 1 hydrogen bond and binding energy of -5.00kcal/mol for PDGFR.
Cyanidin also exhibited inhibition constant above 13.48µm (Figure 2) and
11.64µm (Figure 3) for ERK and RAF. PDGFR have membrane protein tyrosine kinase
and is a therapeutic target in cancer, cardiovascular and autoimmune diseases35.
Based on our insilico study we would like to stress that cyanidin modulates
MAPK kinase pathway by inhibiting PDGFR and modulating EGF, FGF, PDGF, RTK,
RAS, MEK, RAF, ERK, JUN, FOS and SOS. However further insilico and invitro
studies are necessary to validate this claim of ours.
CONCLUSION:
Thus the insilico
docking study establishes the possible role of cyanidin modulating the MAPK
kinase pathway by inhibiting PDGFR and modulating EGF, FGF, PDGF, RTK, RAS,
MEK, RAF, ERK, JUN, FOS and SOS. However further insilico and invitro
studies are necessary to validate its role in the action of cyanidin.
CONFLICT OF INTEREST:
Nil
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