Journal of Receptors and Signal Transduction

ISSN: 1079-9893 (Print) 1532-4281 (Online) Journal homepage: http://www.tandfonline.com/loi/irst20

Zerumbone binding to estrogen receptors: an in-

silico investigation

Eltayeb E. M. Eid, Faizul Azam, Mahmoud Hassan, Ismail M. Taban &

Mohammad A. Halim

To cite this article: Eltayeb E. M. Eid, Faizul Azam, Mahmoud Hassan, Ismail M. Taban &
Mohammad A. Halim (2018): Zerumbone binding to estrogen receptors: an in-silico investigation,
Journal of Receptors and Signal Transduction, DOI: 10.1080/10799893.2018.1531886

To link to this article: https://doi.org/10.1080/10799893.2018.1531886

Published online: 05 Nov 2018.

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JOURNAL OF RECEPTORS AND SIGNAL TRANSDUCTION
https://doi.org/10.1080/10799893.2018.1531886

RESEARCH ARTICLE

Zerumbone binding to estrogen receptors: an in-silico investigation

Eltayeb E. M. Eida, Faizul Azama , Mahmoud Hassanb, Ismail M. Tabanc and Mohammad A. Halimd,e
a
Unaizah College of Pharmacy, Qassim University, Unaizah, Saudi Arabia; bSwiss Tropical & Public Health Institute, University of Basel,
Switzerland; cSchool of Biosciences, Cardiff University, Cardiff, United Kingdom; dDivision of Computer-aided Drug Design, The Red-Green
Research Center, BICCB, Dhaka, Bangladesh; eInstitut Lumi
ere Mati
ere, Universite Lyon 1–CNRS, Universite de Lyon, Villeurbanne
Cedex, France

ABSTRACT ARTICLE HISTORY

Breast cancer is the most frequent malignancy among females worldwide. Estrogen receptor (ER) Received 23 November 2017
mediate important pathophysiological signaling pathways induced by estrogens, and is regarded as a Revised 15 August 2018
promising target for the treatment of breast cancer. Zerumbone (2,6,9,9-tetramethylcycloundeca- Accepted 21 August 2018
2,6,10-trien-1-one; ZER), a chemical constituent present in the Zingiber zerumbet is known to exhibit
KEYWORDS
anti-breast cancer activity by modulating several proteins to induce apoptosis. Medicinal chemists usu- Breast cancer; docking;
ally exploit lead compounds of natural origin to develop molecules with improved pharmacological molecular dynamics;
properties. Current study is intended to utilize molecular modeling techniques to investigate the inter- density functional
action of ZER with estrogen receptors. AutoDock was used to predict the binding modes of ZER and theory; zerumbone
target receptors. Stability of the ZER-ER complex was verified by molecular dynamics simulation using
Desmond software. Docked ZER was further optimized by density functional theory (DFT) using
Gaussian09 program. Analysis of docked conformations in terms of binding energy disclosed estrogen
receptor-b (ERb) as more promising than estrogen receptor-a (ERa). Evaluation of MD trajectories of
ZER bound to both ERa and ERb showed appreciable stability with minimum Ca-atom root mean
square deviation shifts. DFT based global reactivity descriptors such as electron affinity, hardness,
chemical potential, electronegativity and electrophilicity index, calculated from the energies of highest
occupied and lowest unoccupied molecular orbitals underscored the electronic features governing via-
bility of the ZER for interaction with the target receptors. In conclusion, these findings can be
exploited to design and develop novel anticancer agents based on the lead compound, ZER.

Introduction cells, which resulted in suppression of anti-apoptotic and

Natural compounds obtained from plants represent an
immense resource of structurally diverse chemical entities
possessing high affinity and specificity for various pharmaco-
logical targets. They have proven to be by far the magnifi-
cent sources for novel leads in drug discovery and
development programs. In recent times, bioactive molecules
derived from natural sources have served as a privileged
starting point for therapeutic intervention of various human
ailments [1–3].
Zerumbone (2,6,9,9-tetramethylcycloundeca-2,6,10-trien-1-
one; ZER) is a monocyclic sesquiterpene, isolated from the
rhizome of ginger plant (Zingiber zerumbet), found in tropical
and subtropical areas (Figure 1) [4]. ZER is known to exhibit
a diverse range of biological activities including anticancer
activities involving different signaling pathways in various
cell lines [5–7]. It has exhibited promising activity against
breast and other cancers in preclinical models [8–11].
Bounty of preclinical studies involving a variety of cancer
cell types have conferred a wealth of mechanistic insights
into the anticancer effect of ZER. For example: ZER abolished
NF-jB and IjBa kinase activation in a panel of human cancer
metastatic gene expression and, induction of apoptotic cell
death, as well as inhibition of cell invasion [12]; treatment of
cancer cells with ZER modulated Bax/Bcl-2 ratio favoring
apoptosis, inhibited Sonic hedgehog/GLI-mediated transcrip-
tion, and downregulated chemokine receptor CXCR4 con-
comitant with inhibition of CXCL12-induced breast and
pancreatic cancer invasion [7,13,14]. In addition, ZER abol-
ishes RANKL-induced NF-jB activation, inhibits osteoclasto-
genesis, and suppresses human breast cancer-induced bone
loss in athymic nude mice [15].
Estrogens have been recognized as potent mitogens in
the mammary gland in which they regulate the growth,
development, and functioning of normal as well as cancer-
ous breast [16,17]. Role of estrogens in the proliferation and
progression of breast cancer has been further established in
various epidemiological and animal studies where removal of
the ovaries or the treatment with antiestrogens opposes
their deleterious effects [18,19]. Estrogen receptors (ERa and
ERb) belong to a large family of nuclear receptors, which
mediate the genomic action of estrogens by acting as ligand
dependent transcription factors [20,21]. Both of these recep-
tors are encoded by two unique genes and therefore, ERb

CONTACT Eltayeb E.M. Eid eem.eid@qu.edu.sa; Faizul Azam faizulazam@gmail.com, f.azam@qu.edu.sa Unaizah College of Pharmacy, PO Box 5888,
Qassim University, Unaizah-51911, Saudi Arabia
ß 2018 Informa UK Limited, trading as Taylor & Francis Group
2 E. E. M. EID ET AL.
Figure 1. Chemical structure of zerumbone.
shares a high homology to ERa, in particular in the DNA
binding domain (95%) and the ligand binding domain (55%)
[22]. Among these, ERa-positive breast cancer is the most
common type of breast cancer and therefore, depends on
estrogen for growth and development. Consequently, target-
ing estrogen and its receptor is one of the crucial armament-
arium in the therapeutics of breast cancer [23] even though
only 60% of ER-positive tumors respond to adjuvant therapy
with tamoxifen [24]. Tamoxifen is a non-steroidal estrogen
antagonist, considered as drug of choice for the treatment of
ERa-positive breast cancer with both premenopausal and
postmenopausal women [25]. It is a selective estrogen recep-
tor modulator, competes with estrogens in binding to ERs,
blocking the transactivation site of ER leading to the inhib-
ition of estrogen action [26]. Regardless of the beneficial
effects of tamoxifen in the treatment of breast cancer, its
promoter effect on the gynaecological tract may result in the
development of numerous side effects such as hot flashes,
vaginal discharge and bleeding, suppression of menstruation,
and gastrointestinal disturbances [26]. Therefore, novel
potential drug candidates are always required to overcome
the drawbacks of chemotherapeutic agents.
Considering the remarkable anti-breast cancer activity of
ZER, and lack of mechanistic insight into the interactions
with estrogen receptors, current research is envisioned to
illuminate its binding modes with ERa and ERb by employing
molecular docking coupled with molecular dynamics simula-
tion studies. DFT based calculations of ZER is intended to
underscore electronic features governing the ligand-receptor
interactions.
Materials and methods
Molecular docking
Preparation of the receptors
The three-dimensional (3D) crystal structures of ERa (PDB
Code 3ERD) and ERb (PDB Code 1QKM), in complex with
diethylstilbestrol and genistein, respectively, were retrieved
from the Research Collaboratory for Structural Bioinformatics
(RCSB) Protein Data Bank (PDB; http://www.rcsb.org/pdb/
home/home.do). For each crystal structure, the crystallo-
graphic water molecules were removed, the missing resi-
dues/hydrogen atoms were added and the inhibitors from
the crystal structure were used to define the respective
active sites. For MOE docking, macromolecular crystal struc-
tures were protonated by employing Amber99 force field
after removing water molecules.
Preparation of ligand
Structure of the ZER was retrieved from PubChem database.
The structure of the ligand was drawn in ChemBioDraw Ultra
14.0 and converted to its three-dimensional coordinate in
ChemBio3D Ultra 14.0. Geometry optimization was done
using MM2 method of energy minimization and saved in
pdb format. The prepared ligand was used as input file for
AutoDock 4.2 in the next step.
Docking simulation
Molecular docking study was performed by using Lamarckian
genetic algorithm methodology implemented in AutoDock
4.2 program. The standard docking procedure was adopted
for a rigid protein and a flexible ligand (100 independent
runs per ligand). A grid of 60, 60, and 60 points in x, y, and z
directions was built with a grid spacing of 0.375 Å and a dis-
tance-dependent function of the dielectric constant were
used for the calculation of the energetic map. The default
settings were used for all other parameters. In MOE, docking
parameters were set to default values and 100 poses of ZER
were obtained. All docked poses were ranked based on their
binding energies.
Analysis and visualization of docking simulation results
At the end of docking, the best poses were analyzed for
hydrogen bonding or hydrophobic interactions and root
mean square deviation (RMSD) calculations using Discovery
Studio Visualizer 2016 (Accelrys Software Inc.) and PyMol
(The PyMOL Molecular Graphics System) programs. The result
of molecular docking between estrogen receptors and ZER
using AutoDock and MOE programs is summarized in
Table 1.
Molecular dynamics (MD) simulations
In order to inspect the stability of the docked ligand-protein
complexes the 3 D coordinates of the best docking configur-
ation of ZER in complex with ERa and ERb were subjected to
molecular dynamics simulation using Desmond [27,28]. ZER
in complex with ERa and ERb was immersed in orthorhombic
box with 8476 and 8246 water molecules respectively, using
simple point charge (SPC) solvent model and the optimized
potential for liquid simulations (OPLS3) force field [29] was
applied. An appropriate number of Naþ counter-ions were
used to neutralize the complexes. Isothermal-isobaric (NPT)
ensemble class was used with temperature and pressure
adjusted to 300 K and 1.01325 bar, respectively. Simulation
time was set to 5 ns for each system and the coordinates
were saved for every 5 ps. A cutoff radius of 9.0 Å was
applied for short-range van der Waals and Coulomb interac-
tions. Temperature and pressure of the systems were main-
tained by applying Nose–Hoover thermostat [30] and
 JOURNAL OF RECEPTORS AND SIGNAL TRANSDUCTION 3

Table 1. Results obtained after docking of ZER with estrogen receptors.

H-bond interactions Hydrophobic interactions
Program Target DGb (kcal/mol)
a b
RMSD (Å) No. Amino acids No. Amino acids
AutoDock ERa (3ERD) –8.47 1.19 – – 14 Leu-346, Ala-350, Leu-384, Met-388, Leu-391, Phe-
404, Met-421, Ile-424, Phe-425, Leu-428, His-524.
ERb (1QKM) –9.40 0.17 – – 14 Met-295, Leu-298, Ala-302, Met-336, Met-340, Leu-
343, Phe-356, Leu-380, Leu-476
MOE c
ERa (3ERD) –8.34 1.12 – – 15 Leu-346, Thr-347, Ala-350, Trp-383, Leu-384, Met-
388, Leu-391, Phe-404, Met-421, Ile-424, Phe-425,
Leu-428, Gly-521, His-524, Leu-525
ERb (1QKM) –9.25 0.6 – – 13 Met-295, Leu-298, Thr-299, Ala-302, Met-336, Met-
340, Leu-343, Phe-356, Ile-373, Ile-376, Phe-377, Gly-
472, Leu-476
a
Binding free energy.
b
Root mean square deviation.
c
Molecular operating environment.

Figure 2. The validation of accuracy and performance of docking protocol. AutoDock 4.2: The native and docked ligands of ERa, lime green and brown (a); ERb,
cyan and blue (b) respectively. MOE: The native and docked ligands of ERa, pink and yellow (c); ERb, sky blue and red (d) respectively (see colour version of this fig-
ure at www.tandfonline.com/irst).

Martyna–Tobias–Klein [31] methods respectively. The RESPA
integrator was used with a time step of 2.0 fs [32] for the
overall simulations. The systems were minimized and equili-
brated with the default protocols of the Desmond.
Density functional theory (DFT) calculations
Density functional theory (DFT) employing Becke’s three-par-
ameter hybrid model, Lee–Yang–Parr (B3LYP) correlation
functional method at 6–311 G (d,p) level was used to opti-
mize the minimum energy conformation of the docked ZER.
Gaussian09 software program suite [33] was used for these
calculations. DFT based global reactivity descriptors, such as
electron affinity, hardness, softness, chemical potential, elec-
tronegativity, and electrophilicity index were calculated to
inspect the suitability of these descriptors for understanding
the reactive nature and sites of the ZER molecule. The min-
imum energy conformer obtained from molecular docking
study in the form of pdb format, served as input structure
for the DFT calculations.
Results and discussion
Validation of docking protocol
Accuracy of the docking protocol employed in both
AutoDock 4.2 and MOE programs was assessed by redocking
of the native co-crystallized ligands (diethylstilbestrol and
genistein for 3ERD.pdb and 1QKM.pdb respectively) within
the inhibitor binding cavity (IBC) of human ERa and ERb, and
the docked positions were compared to the crystal structure
positions by calculating RMSD values. According to the crite-
ria of validation cited in literature, RMSD values smaller than
2.0 Å indicate that the docking protocol is capable of accur-
ately predicting the binding orientation of the co-crystallized
ligand [34]. In the present study, RMSD values of both ERa
and ERb were within 2.0 Å (Figure 2), which signifies that our
4 E. E. M. EID ET AL.

Figure 3. Molecular docking results employing AutoDock 4.2. Three dimensional structures of proteins showing the binding sites (left), and main residues involved
in the ligand-protein interactions (right) of ZER with ERa (a) and ERb (b). Both proteins are shown as ribbon style carrying docked ZER as mesh in dark pink color.
Residues of binding pocket of ERa are shown as green sticks and docked ZER as purple ball and stick style (a). ERb binding pocket is presented as stick in cyan
color and docked ZER as ball and stick in red color (b). Ligand-protein interactions are marked by broken lines in purple color (see colour version of this figure at
www.tandfonline.com/irst).

docking methods are valid for the given structures and both
AutoDock 4.2 and MOE, therefore can be relied for docking
ZER into the IBC of ERa and ERb [35–38].
Binding interactions of ZER with estrogen receptors
Estrogen hormones, in addition to regulating multiple
physiological processes, also play a crucial role in promoting
the proliferation of the neoplastic breast epithelium by act-
ing on soluble nuclear estrogen receptors, ERa and ERb [39].
Molecular docking has been routinely employed for several
purposes including ligand binding affinity prediction, ligand
pose prediction and lead identification [3,35,40,41].
Therefore, this methodology can be exploited for the identifi-
cation of novel anti-breast cancer lead by targeting estrogen
receptors, owing to its critical role in gene expression and
transcription [42]. The X-ray coordinates of ERa (pdb id:
3ERD) and ERb (pdb id: 1QKM) co-crystallized with diethylstil-
bestrol and genistein respectively, were downloaded from
the protein data bank website (www.rcsb.org), and processed
further for docking simulations employing AutoDock and
MOE programs. After successful completion of molecular
docking of ZER with each target protein, a total of 100 poses
belonging to docked ZER were visualized to identify the
conformation having minimum binding energy and best lig-
and-receptor interactions. The results in terms of binding
free energy, RMSD value and number of interactions are pre-
sented in Table 1.
There are twelve a-helices (H1–H12) and a b-hairpin in the
ligand binding domain of estrogen receptors. However, H12
is regarded as an indispensable helix for receptor activation
and acts as switch of ligand-binding domain, by adopting a
characteristic conformation upon ligand binding [43,44]. The
ligand-binding domain of ERb differs at only two amino acid
positions to that of ERa. However, overall smaller size of the
ERb pocket as well as amino acid differences of the ligand-
binding pocket may contribute to the identification of com-
pounds displaying receptor selectivity. Unfortunately, ligands
having appreciable selectivity are yet to be identified and cur-
rently studied ligands only display modest selectivity for ERb
versus ERa and certainly there is a necessity to develop novel
compounds with improved selectivity [45].
The molecular docking results by AutoDock showed that
ZER possesses a very similar binding mode as native co-crystal-
lized ligand, diethylstilbesterol by occupying the entire hydro-
phobic pocket of ligand binding domain in ERa (Figure 3).
Common residues involving hydrophobic interactions include
Leu346, Ala350, Leu384, Phe404, Met421, Gly521, Leu525.

Figure 4. Molecular docking results employing MOE. Three-dimensional structures of proteins showing the binding sites (left), and two-dimensional view of main
residues involved in the ligand-protein interactions (right) of ZER with ERa (a) and ERb (b). Docked ZER in both receptors is shown as stick style in green color (see
colour version of this figure at www.tandfonline.com/irst).
However, Thr347, Met388, Ile424, Phe425, and Leu428 were
observed to anchor additional hydrophobic contact in docked
ZER. Likewise, docked ZER also occupies the binding pocket of
ERb in similar fashion to the native co-crystallized ligand, gen-
istein and shares hydrophobic contacts through Met295,
Leu298, Leu339, Met340, Phe356, Ile376, and Leu476 (Figure
3). Other residues encircling the docked ZER by hydrophobic
links in ERb include Thr299, Ala302, Met336, Leu343, and
Leu380. However, keto group, as one and only polar moiety of
ZER could not avail the opportunity to participate in any kind
of hydrophilic interaction within the active sites of both ERa
and ERb. Similarly, results of molecular docking studies by
MOE correspond with AutoDock results, revealing better inter-
action profile of ZER with the ERb in comparison with ERa,
JOURNAL OF RECEPTORS AND SIGNAL TRANSDUCTION 5
exhibiting binding energies of 9.25 and 8.34 Kcal mol1
respectively, as shown in Table 1. All of the intermolecular
interactions were of hydrophobic type while none of the
amino acid residues were observed to interact with the only
hydrophilic keto moiety of the ZER. Most of the interacting
residues of ERa and ERb were common in both AutoDock and
MOE results. However, Thr-347, Trp-383, Gly-521, and Leu-525
residues were found to contribute in the hydrophobic inter-
action within ERa binding site but not observed in case of
AutoDock (Figure 4). Furthermore, Thr-299, Ile-373, Ile-376,
Phe-377, and Gly-472 were also part of the hydrophobic
pocket of ERb receptor but not in case of AutoDock.
Interestingly, Leu-380 residue encircled the binding cavity of
6 E. E. M. EID ET AL.

ERb in AutoDock but MOE result was devoid of this

amino acid.

Molecular dynamics (MD) simulation study of ZER and

estrogen receptors
The molecular dynamics simulations studies were performed
for 5 ns to analyze the steadiness and conformational stabil-
ity of ZER–ERa and ZER–ERb complexes, which were
embedded with water molecules in the physiological envir-
onmental conditions, such as temperature and pressure. The
RMSDs for the Ca of the receptor (in blue) and for the ligand
(in red) are presented in Figure 5. The 5 ns simulation studies
indicate that ZER forms a stable complex with both ERa and
ERb receptors. RMSD values with respect to the progression
of time are often used to inspect whether or not a simulated
system attains stability [46]. RMSD values of protein, ranging
from 0.6–1.8 Å and 0.75–2.25 Å for ERa and ERb, respectively,
is based on all backbone Ca atoms relative to the corre-
sponding starting structures of all trajectories for the simu-
lated ZER-ER complex. RMSD values of ZER shown on right
Figure 5. The Root Mean Square Deviations (RMSD) of backbone atoms relative
Y-axis of Figure 5 specifies how stable the ZER is with
to the starting complexes during 5 ns MD simulation of ZER in complex with
respect to the estrogen receptors and their binding pocket. ERa (a) ERb (b). Each plot shows the RMSD of protein on left Y-axis whereas lig-
The plots of both Ca and ‘Lig fit Prot’ (ligand fitting on pro- and RMSD is presented on right Y-axis (see colour version of this figure at
www.tandfonline.com/irst).
tein) shows the RMSD of ZER when the protein-ligand com-
plex is first aligned on the protein backbone of the reference
and then the RMSD of the ligand heavy atoms is measured.
In both ERa and ERb simulated systems, the RMSD values
increased quickly during the initial time of simulation and
then reached plateaus at nearly 0.5 nsec (ERa) and 1.0 nsec
(ERb), which simply implicate that the simulated systems
achieved stability.
Figure 6 portrays the plot of root-mean-square fluctua-
tions (RMSF) in the orientation of side chain of each residue
and backbone of the protein in the MD trajectory. RMSF for
ERa was observed to be within 2.8 Å whereas in case of
ERb, the fluctuations were recorded up to 4.0 Å. Therefore,
it can be implied that the lower atomic fluctuation for resi-
dues and its backbone atoms is due to small conform-
ational changes during the simulation of both ERa and ERb
receptors [47].
At the end of the simulation, each system was independ-
ently scrutinized for per-residue interaction analysis, which is
based on the average value of the interaction occupancies
of the binding site residues and the results were described
in the form of stacked bar plots, normalized over the course
of the trajectory (Figure 7).
In the case of ZER-ERa system, maximum interaction occu-
pancy was noted for Ala350 where hydrophobic interaction Figure 6. The Root Mean Square Fluctuation (RMSF) of complexes during 5 ns
was maintained during approx. 30% of the simulation time. MD simulation of ERa (a) and ERb (b) (see colour version of this figure at www.
tandfonline.com/irst).
However, ZER-ERb system was demonstrated to preserve
similar type of interaction by Ile373, during approx. 20% of
the simulation time. Amino acids such as Leu346, Ala350,
Leu384, Met388, and Leu525 showing hydrophobic interac- crystallized ligand, diethylstilbestrol in ERa. However, add-
tions in docked ZER were also observed to sustain their par- itional residues including Met343, Leu349, Leu387, Leu391,
ticipation in such interactions throughout the simulated Phe404, Met421, Ile424, Phe425, Leu428, Val534, and Leu540
trajectory of ZER–ERa complex. Similar residues are also were also observed to play significant role in hydrophobic
known to constitute the binding pocket of native co- interactions (Figure 8). Similarly, residues Met295, Leu298,

Figure 7. Per-residues analysis of the ZER in complex with ERa (a) and ERb (b). The analysis was based throughout the 5-ns MD simulations. Hydrogen bond,
hydrophobic and water bridge interactions are illustrated by green, purple and blue lines, respectively (see colour version of this figure at www.tandfonline.
com/irst).
Leu339, Met340, Phe356, Ile376, and Leu476 showing their
importance in hydrophobic interactions of docked ZER also
conserved their contribution in simulated ZER-ERb system.
However, it was interesting to note that these residues also
outline the binding pocket of native co-crystallized ligand,
genistein, in ERb.
DFT computations
Molecular interactions of ligands with target proteins are
absolutely crucial to various pharmacological activities, which
in turn depend on the structural features of the molecules.
Density functional theory (DFT) calculations furnish informa-
tion about electronic effects and intermolecular charge trans-
fer responsible for ligand-protein connections [48–50].
Therefore, computation of the HOMO (highest occupied
molecular orbital) and LUMO (lowest unoccupied molecular
orbital) energies describes the reactivity, shape and binding
properties of a complete molecule as well as of molecular
fragments and substituents. In recent times, the concept of
HOMO and LUMO has been successfully executed in explicat-
ing the biological activity and molecular properties of the
drug candidates [47,51,52].
JOURNAL OF RECEPTORS AND SIGNAL TRANSDUCTION 7
The energy separation between frontier molecular orbitals
constituting the gap between the energies of highest occu-
pied molecular orbital (HOMO) and lowest unoccupied
molecular orbital (LUMO) reflects the kinetic stability and
chemical reactivity of the molecule [47,52,53]. The HOMO
and LUMO energy gap for ZER was calculated at the B3LYP/
6-311G(d,p) level and the graphical representation of these
orbitals is shown in Figure 9. The positive and negative
phases of molecular orbitals can be recognized in red and
green color respectively. Detailed inspection of HOMO and
LUMO orbitals indicates that the path of electron density
transfer from ground to first excited state. Both of these are
considered as key orbitals contributing in chemical reactivity
as well as bioactivity. In HOMO, the electron distribution is
scattered over the cross conjugated ketone moiety covering
whole a,b-unsaturated carbonyl group, and partially extend-
ing towards C8 as well as C15 in the 11-membered ring
structure of ZER. However, LUMO has been detected to par-
tially spread on C13 and C9 in addition to the carbonyl moi-
ety and both conjugated double bonds. Interestingly,
isolated double bond between C10 and C12 conferred not
any participation in HOMO/LUMO transition. According to
Koopmans’ theorem [54] the ionization potential (I) and elec-
tron affinity (A) of ZER can be expressed through HOMO and
8 E. E. M. EID ET AL.

Figure 8. Ligplot diagrams of docked ZER into the binding cavity of ERa (a) and ERb (c). A representation of detailed ligand atom interactions during MD simula-
tion of ZER with the protein residues of ERa (b) and ERb (d) (see colour version of this figure at www.tandfonline.com/irst).

LUMO orbital energies as: I ¼ EHOMO ¼ 6.46 eV and Conclusion

A ¼ ELUMO ¼ 1.33 and the HOMO-LUMO gap ¼ 5.13 eV. Based
on this, chemical potential, l ¼ ðELUMO þ2 EHOMOÞ ¼ 3.90 eV
and hardness, g ¼ ELUMO2 EHOMO ¼ 2.56 eV were computed. A
large HOMO/LUMO energy gap is associated with a hard
molecule and small HOMO/LUMO gap with a soft molecule.
So, a molecule with smaller HOMO/LUMO gap is expected to
be more reactive. Electrophilicity index (x) of ZER was calcu-
lated by implementing the formula of [55] as
l2
x ¼ 2g ¼ 2.96 eV, which defines a quantitative classification of
global electrophilic nature of a compound. They anticipated
x as a measure of energy lowering due to maximal electron
flow between donor and acceptor.
Molecular interaction of ZER and estrogen receptors was
studied by employing docking, molecular dynamics simula-
tion and DFT based optimization. Docking results showed
appreciable affinities with both ERa and ERb receptors and
demonstrated similar binding interactions with native co-
crystallized ligands of the respective receptors exhibiting
exclusively hydrophobic interactions. However, keto group,
as one and only polar moiety of ZER could not afford any
kind of hydrophilic interaction within the active sites of
both estrogen receptor types. Results of MD simulation of
docked-ERa/ERb systems in terms of RMSD, RMSF and per-

Figure 9. DFT optimized structure of ZER (a) generated at B3LYP/6-311G (d,p). Visualization of LUMO (lowest unoccupied molecular orbital; (b) and HOMO (highly
occupied molecular orbital; (c) (see colour version of this figure at www.tandfonline.com/irst).
residue interaction analyses authenticate the stability of the
studied complexes throughout the simulated trajectories.
DFT-based global reactivity descriptors, such as electron
affinity, hardness, chemical potential, electronegativity, and
electrophilicity index can be implemented to better under-
stand the ligand-receptor interactions of ZER. These results
provide an insight into the design and development of novel
anticancer agents based on the lead compound, ZER.
Disclosure statement
No potential conflict of interest was reported by the authors
ORCID
Faizul Azam http://orcid.org/0000-0002-2927-8167
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