andiblthcaTorereangecodrofenre
St
wanblis a sti
Gy66e-m
ScGasu
Gy66
Gy66
NaRe
NaRe
proGa+8
H
Abstract—Hnd blood vesselseases. The Reood pressure coe rate-limiting s
an inhibit renin o achieve thceptor-based phnnin-inhibitor c
nd scaffolds oenerated considompounds mapprug-like screeninf renin. Finallynergy was selecnin inhibitors. Keywords—R
tructure-based p
YPERTENassociated
worldwide [1]. nd often requilood pressure an important key role in thimulated by a
Chanin Park is wyeongsang Nation60-701, Republic mail: [email protected]
cience, Gyeongazwa-dong, Ji
[email protected] Lee is wi
yeongsang Nation60-701, Republic
Minky Son is wyeongsang Nation60-701, Republic
Shalini John is ational Universitepublic of Korea
Young-sik Sohnational Universitepublic of Korea
Keun Woo Leeogram), Gyeongazwa-dong, Jinju 82-55-772-1359;
MolecPha
Chanin P
H
Hypertension is ls thus increasi
enin angiotensinontrol. Renin isstep. Our aim isand thereby em
his, molecular harmacophore mcomplex structuof inhibitors. Tdering conformped to these mng. The identifi
y, hit1 satisfyincted as possibl
Renin inhibitopharmacophore
I. INT
NSION is a d with signi
It remains pires combinatilowering. Thregulator of s
he control of ba number of s
with the Divisionnal University (Gof Korea (phone
gnu.ac.kr). nThangapandian sang National inju 660-701,c.kr). ith the Division onal University (Gof Korea (e-mail
with the Division nal University (Gof Korea (e-mailwith the Divisio
ty (GNU), 501 (e-mail: shalini@
n is with the Divity (GNU), 501 (e-mail: ysohn@e is with the Dgsang National 660-701, Republe-mail: kwlee@b
cular Darmac
DPark, Sundar
characterized wing the risk of n system (RASs the enzyme ths to develop new
merge as therapedynamics (M
modeling were ures were selectThree pharmacmations induc
models were selied hits were dong the binding le lead candida
or, Molecular modeling.
RODUCTION major cardioficant morbioorly controllion therapy toe renin-angiotsalt and water blood pressuresignals, includ
n of Applied Life GNU), 501 Jinju-: +82-55-772-136
is with the DiUniversity (G
, Republic
of Applied Life GNU), 501 Jinju-l: [email protected] Applied Life
GNU), 501 Jinju-l: [email protected] of Applied LJinju-daero, Gaz
@bio.gnu.ac.kr). ision of Applied LJinju-daero, Gaz
@bio.gnu.ac.kr). Division of Appl
University (Glic of Korea (phonbio.gnu.ac.kr).
DynamcophorDiscorapandianTh
with stress on tf heart attack anS) plays a majohat controls thew drug-like leadeutics for hyperMD) simulatiimplemented, ated based on ICcophore modeed by inhibitlected and subj
ocked into the acmode and int
ate to be used i
dynamics sim
ovascular riskidity and mled in many p
o achieve the ttensin systemhomeostasis a
e (Fig. 1).The ding a drop in
Science (BK21 p-daero, Gazwa-do60; fax: +82-55-7
ivision of ApplGNU), 501 Jin
of Korea
Science (BK21 p-daero, Gazwa-doac.kr). Science (BK21 p
-daero, Gazwa-dou.ac.kr).
Life Science, Gyezwa-dong, Jinju
Life Science, Gyzwa-dong, Jinju
lied Life SciencGNU), 501 Jinne: +82-55-772-1
mic Sire Moovery ohangapandian
the heart nd renal r role in
e RAS at ds which rtension. on and
and three C50 value els were or. The jected to ctive site teraction in novel
mulation,
k factor mortality
patients targeted
m (RAS) and has RAS is n blood
program), ong, Jinju 772-1359;
lied Life nju-daero,
(e-mail:
program), ong, Jinju
program), ong, Jinju
eongsang 660-701,
eongsang 660-701,
ce (BK21 nju-daero, 1360; fax:
preplaeffanforThanThrecultrenrenan
asp2 lcleresanrencatan[9]
Fig
wianhasubfoltreof thetha
imulatdelingof Novn, Yuno Lee,Keun Woo L
essure, a decrasma sodium fective way to
nd renal disordrm the hemodhe active octngiotensin I bhe binding ofceptors (AT1timately leadinin controls thnin inhibitio
ntihypertensiveThe human rpartyl proteaselobes with a left, the activesidue of angio
nd Asp226 in nin, control thtalyzing the h
ngiotensinogen].
g. 1 The Renin ARenin is
Renin inhibitith fewer side
ntagonists [10]ave started wibstrate angiotllowed by vareating hypertenf these compouere was a needat fulfill all
tion ang on Hvel In Minky Son,Lee
rease in circuconcentratio
o intervene in tders [2]. The dynamically intapeptide ang
by the angiotef angiotensin ) triggers a ning to an incrhe first rate-lion is conse strategy [4]-renin, a 340-e superfamily long and deepe site of reninotensinogen. Thuman renin)he first and r
hydrolysis of tn and releasin
Angiotensin Sys the rate-limiti
ors have beeneffects than A. The design a
ith peptide miensinogen. Thrious syntheticnsion as reninunds survivedd for new clascriteria for
nd ReHumanhibito, Shalini John
ulating volumon. Inhibition the pathogenerenin cleaves
nactive decapegiotensin II ensin-converti
II to the tynumber of phrease in bloodimiting step isidered to -[5]. amino acid, i [6]. Structurap cleft between, accommodaTwo aspartic a), each locatedrate-limiting sthe Leu10-Vang the decape
ystem (RAS) aning step in the R
n predicted to ACE inhibitorand developmeimicking the hese peptide bc peptides we
n inhibitors [8]d the stages ofsses of nonpepbecoming su
ecepton Reniors
n, Young-sik
me, or a reducof the RAS
sis of cardiovas angiotensinoeptide angioteis converted
ing enzyme (ype-1 angiotenhysiological ed pressure [3]in the RAS ca
be an att
is a member ally, renin conen them [7]-[8ates a 7-aminacid residues (d in a single lstep of the Ral11 peptide beptide Angiote
d the sites of bl
RAS cascade
be more efficrs and AT1 reent of renin infeatures of itased renin inh
ere not succes], [11]-[13]. Af drug developtide renin inhuccessful drug
or-basein for
k Sohn, and
tion in S is an ascular ogen to ensin I. d from (ACE). nsin II effects, ].Since ascade, tractive
of the sists of 8]. The no acid (Asp38 lobe of
RAS by bond of ensin I
ockade.
cacious eceptor nhibitor ts only hibitors ssful in
As none pment,
hibitors gs [8].
ed
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Among the various efforts taken to develop renin inhibitors with good bioavailability, only aliskiren arose from the former peptidomimetic structures as a successful candidate. Aliskiren is the only renin inhibitor launched in the market as anti-hypertensive agent [14].
In this study, three receptor-based pharmacophore models were developed to consider the conformational diversity of the active site depending on bound. The MD simulations in general consider the conformational dynamics of protein by generating an ensemble of protein conformations. Developing a pharmacophore model taking into account the conformational changes in the active site is rational and will increase the reliability of the models [16]. These pharmacophore models were used in database screening to identify new and potential hits for future drug design. Molecular docking study has reduced the probability of picking false positives as potential hits. This methodology will be useful in identifying new renin inhibitors with similar binding orientation as existing compounds.
II. METHODS A. Preparation of Systems Among dozens of x-ray crystal structures of human
renin-inhibitor complexes available in the protein data bank (PDB), three complex structures (PDB ID: 3G6Z [15], 3GW5 [17], and 2V0Z [8]) were selected based on the IC50 value (< 1.0 nM) and chemical scaffolds of bound inhibitors. These selected crystal structures have contained missing region at Glu167-Gln170, Leu1-Leu3, and Asn168, respectively. The complete structures that were repaired the missing region were prepared through the Build Homology Models protocol within Discovery Studio (DS) 2.5.
B. Molecular Dynamics Simulation The MD simulations of three renin-inhibitor complex
structures were performed with GROMOS96 force field using GROMACS 4.0.7 package [18]. Initially, all the ionizable residues in the protein were protonated at pH7. The topologies and charges for each inhibitor were calculated using the PRODRG web-server [19]. Each complex was solvated with the SPC water molecules. Eight Na+ counter-ions were addedto ensure the overall charge neutrality of the system. Energy minimization was performed with steepest descent algorithm. Then the protein was restrained and the solvent molecules with counter-ions were allowed to move during a 100 ps position-restrained MD run. The well equilibrated structure was then used for 5 ns production runs. The electrostatic contributions were calculated using the particle-mesh Ewald (PME) algorithm [20] with a direct interaction cut-off of 0.9 nm and grid spacing of 0.12 nm. The van der Waals (VDW) forces were treated by using a cut-off of 1.4 nm. All simulations were executed under periodic boundary conditions with the NPT ensemble using the V-rescale thermostat and Parrinello-Rahmanbarostat for keeping the temperature (300 K) and the pressure (1 bar) constant. Bonds between heavy atoms and corresponding hydrogen atoms were constrained to their equilibrium bond lengths using the LINCS algorithm [21] and
the geometry of water molecules was constrained using the SETTLE algorithm [22]. The time step for the simulation was 2 fs and the coordinates were stored every 1 ps.
C. Cluster Analysis Selection of a representative structure which could represent
the conformational flexibility of the protein throughout the simulation time is crucial. Therefore, three representative structures were selected from each simulation trajectory of 5000 frames saved during the simulation time. The g_cluster implemented in GROMACS was used for cluster analysis of the trajectories from 2001 ps to 5000 ps. From the RMSD comparisons of each snapshot with all the others, clusters were generated with the cut-off values of 0.143 nm (3G6Z), 0.128 nm (3GW5), and 0.120 nm (2V0Z) using gromos method.
D. Receptor-Based Pharmacophore Model Generaion The three representative structures obtained from the most
populated clusters were used to generate receptor-based pharmacophore models screening the complementary pharmacophoric features in the active site of protein. The Interaction Generation protocol in DS was used to extract all the available hydrophilic and hydrophobic interaction points that can be complemented by the bound inhibitor. The representative features including hydrogen bond acceptor (HBA), hydrogen bond donor (HBD), and hydrophobic (HYP) features were selected based on the active site residues. The final pharmacophore model consists of the essential pharmacophoric features, which are supposed to be present in the inhibitor.
E. Database Screening and Drug-Like Filtration The three receptor-based pharmacophore models were used
as 3D structural queries in database screening in order to find potential hit compounds suitable for further progress. The screening calculation was carried out using the Ligand Pharmacophore Mapping protocol with the Best/Flexible search option in DS. Two chemical databases containing diverse chemical compounds were used in database screening. The fit value for filtering hit compounds was set based on the fit values of already known renin inhibitors. Hit compounds with higher fit value than any of the fit value of known most active compounds were selected and checked for their drug-like properties using Lipinski’s rule of five and ADMET filters using DS. Final hit compounds that passed all of these screening tests were selected and used in molecular docking study.
F. Molecular Docking and Interaction Energy Molecular docking studies for final hit compounds were
performed using GOLD 4.1 program [23]. The active sites of three representative structures were defined with a radius of 10 Å around the bound inhibitor. All hit compounds from database screening along with the bound inhibitors in three crystal structures were docked into the active site. The Maximum save conformations was set to 10 and the Early Termination was set as 5 to skip the calculation if the RMSD between any of the 5 docked conformations is less than 1.5 Å. Energy minimization
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cafowovan5,DInusIm
codestrcoinstr3GwC4inlocoacstrorM
Rrerethronu
wfusimvathRre
alculations weor three system
was used to relverlaps that prnd protein. T,000 max steistance-Depen
nteraction enersing the Calmplicit Distanc
A. Selection aThree renin-
onsider the epending on ructures 3G6Zompounds wernhibitors, respructurally divG6Z contains
whereas the 72X41 (Inh3) of
nhibitor, whichong substitutonformationalccommodate dructures withrders, and add
MD simulation
LIST OF CRYSTA
PDB ID
Re
3G6Z
3GW5
2V0Z
B. Stability ofThree MD
enin-Inh3 comefined and eceptor-based hree simulatioot-mean-squaumber of intra
The RMSD vwere measured unction of the mulations wealues betweenhe simulationenin-Inh3 sys
espectively. A
ere performedms using DS.ax the dockedroduce bad conThe parametereps, RMS Grndent Dielectrgies for all miculate Interacce-Dependent
III. and Preparati-inhibitor comconformationthe bound
Z, 3GW5, andre 0.16nM, 0.4pectively. Th
verse scaffolds a fused ringX (Inh2) of 3Gf 2V0Z is alh contains onltions. Thesel diversity ofdiverse chemih recovered mded hydrogensn studies.
TAL STRUCTURES O
esolution (Å)
2.00
2.00
2.20
f Simulations simulations mplex structuadjusted strupharmacopho
ions were are deviation (a-protein hydrvalues of the pwith respect simulation timre achieved in
n 0.15 nm and n, the RMSstems increaseAfter this tim
d to refine top. The Smart Md conformationtacts betweenrs for energyradient of 0.1trics for impinimized strucction Energyt Dielectrics m
RESULTS ion of Structurmplex structurnal diversity
inhibitors (Td 2V0Z, the I4nM, and 0.6nhese bound s to each otherg system withGW5 is a piperiskiren, the fly one substitue three struf active site ical compoundmissing regio were taken as
TABLE I OF RENIN WITH D
Inhibitor
of Renin-Inhures were peructures usedore models. Texamined b
(RMSD), poterogen bonds. protein Cα atoto their initialme (Fig. 2a). n the atom po0.3 nm. Durin
SD values oed up to ~ 0.1me, the valu
p 5 docked strMinimizer algns and removn docked com
y minimizatio1 kcal/mol, a
plicit solvent ctures were mey protocol wmodel option i
res res were seleof the activ
Table I). ForIC50 values ofnM as the moscompounds
r. The A7T (Ih other substiridine derivatifirst marketeduted phenyl rinuctures refle
of renin thds. These threons, correcteds initial structu
DIFFERENT INHIBI
IC(n
A7T (Inh1) 0
72X (Inh2) 0
C41 (Inh3) 0
h1, Renin-Inhrformed to cod to generaThe stabilitiesby calculatinential energy,
oms for three sl configuratioThe stabilitie
ositional RMSng the first 60of Renin-Inh17 nm and ~ 0es of RMSD
ructures gorithm ve steric
mpounds on used and the model.
easured with the in DS.
ected to ve site r three f bound st active are of
Inh1) of tutions,
ive. The d renin ng with
ect the hat can ee x-ray d bond ures for
ITORS C50 nM)
.16
.47
0.6
h2, and onstruct ate the s of the ng the and the
systems ns, as a s of the
SD with 00 ps of h1 and 0.2 nm, D were
mahagra20cansim
intouen(RkJ/enkeptheabtheboThpacowerel
FigRMc t
recpe3).coIn ch(2coclusna39strstrrestherepph
aintained untiand, the RMadually increa
000 ps. From tn conclude mulation envirThe calculatitra-protein hy
ut to confirm tnergy of each sRenin-Inh1), -
/mol (Renin-Inergetically stapt about 225 e simulation (normal confoe simulation. A
onds was simihese differencattern betweennclusion, thesere stable duriliability with n
g. 2 Results of MSD with respethe number of i
C. Selection ofIn order to sceptor-based prformed using. The conformmparing the aRenin-Inh1
haracterized an155 snapshotsnformations, uster among napshots) of th
961 ps (Rep2),ructures of eaructures of spectively. The dynamic conpresentative s
harmacophore
il the end of tSD value ofase up to ~ 0.2the RMSD anthat the threronment durinions of potendrogen bondsthe stabilities system was ma
760559 kJ/mInh3), which able (Fig. 2b).intra-protein
(Fig. 2c). Theormational chaAlthough the ilar, potentialces appear ton active site rse analyses haing the simulatno artifacts.
simulation stabect to the initialintra-protein hy
the 5 ns M
of Representatselect the reprpharmacophog the snapshot
mational familactive site con
and Renin-Innd the top clus) and 61.47 respectively.
nine clusters whe total confo, and 3878 ps
ach top clusteRenin-Inh1,
hese structuresnformational structures wer
models comp
the simulationf Renin-Inh2 28 nm and kepnalysis of the Mee systems wng the simulatntial energies s of the system
of the simulaaintained at abmol (Renin-Inindicates that. In addition, hydrogen bon
ese results shoanges in the number of intl energies difo result from residues and bave revealed thtion time and
bility analyses fl structure, b theydrogen bonds aMD simulation
tive Structuresresentative strre models, clts obtained froies were statisformations ofnh3, totally
usters were co% (1844 snaIn case of R
was compriseformations. Th
(Rep3) snapsr were select
Renin-Inh2,s would be appspace of the a
re utilized in plementing th
n time. On thewas continu
pt a stable leveMD simulatiowere stable ion time. and the num
ms were also ations. The pobout - 833828 nh2), and - 7t the simulatioall the systemnds for the peow that there proteins throutra-protein hyffer in each s
diverse interbound inhibithat all three sythus confirme
for three systeme potential enerare plotted throu
s ructures to geluster analyseom last 3000 pstically classiff the 3000 snapeight clusters
omposed of 71apshots) of thRenin-Inh2, td of 55.83 %he 3667 ps (shots that are ed as represe and Renin
propriate to repactive site. Althe developm
he active site
e other ued to el after
ons, we in the
mber of carried otential kJ/mol
798930 ons are
ms were riod of are no ughout drogen system. raction tors. In ystems ed their
ms. a Cα rgy, and ughout
enerate es were ps (Fig. fied by pshots. s were 1.83 % he total the top
% (1675 (Rep1), middle
entative n-Inh3, present ll three
ment of of the
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pr
Fi
sycain
FiB
aloar
berethinInTybohysyhy
rotein.
ig. 3 The histog
To investigatystems, comparried out thronhibitors with
ig. 4 Binding moBinding conforong with hydroare colored in bresidues are sho
CONS
Interactions
Hydrogen bond
interaction
Hydrophobic
interaction
In Rep1, onletween Inh1 esidue, Tyr20,hrough hydrognteractions wenh3 has formedyr20, Gly40, aond interactionydrogen bondystems, Tyr20ydrogen bond
gram obtained frthe 5 n
te the conservparison of through analyzinthe active site
ode of each inhrmations of a Inogen bond interablue, green, and own in stick mo
represented b
TERVED RESIDUES
Rep1
d Ser230
Gln19, V
Leu121
ly one hydrogand Ser230 (, Thr85, and gen bonding
ere observed bd a hydrogen band Thr309 (Fns between Ind interacting 0 and Ser84 ing in both Re
from the cluster ns simulation
ved interactingree representa
ng the interacte residues (Fig
hibitor in three renh1, b Inh2, andacting residues.
d red, respectiveodel. Hydrogen by green dotted
ABLE II
S INVOLVED IN IN
Rep2 Tyr20
Ser84
Thr85
Ser230
al36, Trp45, Tyr8
, Ala122, Phe124
gen bond inte(Fig. 4a). In Ser230, has (Fig. 4b). T
between Inh2 bond interactiFig. 4c). Therenh3 and Ser84.
residue conwere found
ep2 and Rep3
analyses of las
g residues in aative structurtion patterns og. 4).
epresentative std c Inh3 are com. Rep1, Rep2, anely. The inhibito
bond interactioline
NTERACTIONS Rep3
0
Tyr2
Gly4
Ser8
Thr30
83, Pro118, Phe1
4, Val127, Ala22
eraction was case of Rep2interacted wit
Two hydrogenand Ser84. Inon with each re were two hy. Although thenserved in ald to be invol3, and Ser230
st 3 ns of
all three res was of three
tructures. mpared nd Rep3 ors and ons are
3 20
40
4
09
19,
9
formed 2, each th Inh2 n bond n Rep3, residue, ydrogen ere is no ll three lved in in both
Reintobstrresph
usiangefeadiract
F
cHBre
an(PhgeSecoTyinhgeresanphusiPhwecoweThchthecarcarduph
ep1 and Rep2teracting resid
bserved. Fromructures, key sidues were
harmacophore
D. Receptor-BThree recepting the repre
nalyses. The Hnerated basedature, but sperection in whitive site residu
Fig. 5 The recepthree represent
Pharm1, b Phconstraints are cBA, HBD, and Hespectively. Res
green, and
Six pharmacond three HYPharm1) from nerated as co
er84 and Tymplimentary
yr20 and Ser84hibitor duringnerated to hsidues and wo
nd HBD featurharmacophore ing Rep2 and
harm1, the phere also createmponents (Fiere also made he HBA and hosen based one HBD featurerboxyl grouprboxyl group
uring simulaharmacophore
2 (Table II). Idues conservem the comp
hydrogen boidentified anmodels.
Based Pharmator-based phasentative stru
HBA, HBD ad on the consiecific featuresich each inhibues of renin (F
ptor-based phartative structuresharm2, and c Phcomplemented hHYP features arsidues of Rep1,d red, respective
ophoric featureP were selec
Rep1. Two omplementaryr20. A HBDfeature to the
4 involve in thg the simulatiohydrophobicallorked as the pres (Fig. 5a). models, PharRep3, respectarmacophoriced complemenig. 5b, c). Thof two HBA, HYP feature
n the same resie was selectedp of Asp226
of Asp38 mation time.
models revea
In addition, toed in all threeparison of tond and hydrnd then cons
acophore Modarmacophore uctures obtainand HYP chemideration of ths were chosenbitor formed iFig. 5).
rmacophore mos. 3D pharmacoharm3 with intehydrogen bond re shown in gre Rep2, and Repely, and shown
es containing ted as a phaHBA feature
y features to D feature we carboxyl grhe hydrogen bon. The three ly interact w
proper connectIn addition to
rm2 and Pharmtively. Similarc features of Pntary to the ihese two phaone HBD, and
es of Pharm2idues as in Phad as a complim
instead of Amoved aside f
Comparison aled that each m
otal 11 hydroe systems werhree represerophobic intersidered to ge
del Generationmodels were
ned from the mical featurehe direction on depending interactions w
odels generated ophore features er-feature distand interacting resen, magenta, an
p3 are colored in in stick model
two HBA, onarmacophorees of Pharm1
the main chas generatedroup of Asp3bond interactio
HYP featurewith the activting points foo Pharm1, twom3, were devr to the generaPharm2 and Pimportant actiarmacophore md three HYP fe2 and Pharm3arm1, but in Ph
mentary featureAsp38. Becaufrom the activ
of these model contain
ophobic re also
entative racting enerate
n e built cluster s were
of each on the
with the
from of a
nce idues.
nd cyan, n blue,
e HBD, model
1 were hain of d as a 8. The
on with es were ve site r HBA o more
veloped ation of Pharm3 ive site models
features. 3 were harm3, e to the use the ve site
three ns same
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phdich
3DneAtothAcosh
reFithOfrowco
asprsitseorThsucoenanIII
harmacophoriistance constrhanges upon in
E. Database The three dev
D structural qew renin inh
Asinex and Mao the pharmacheir drug-like
ADMET filterompounds retrhown in Fig. 6
Fig. 6 Datab
From Asinexetrieved througinally, 11 com
hree pharmacon the other haom Maybridg
were obtainedonsidered for m
F. MolecularThe molecu
ssess the 12 hirocess and to ite of renin. Fr
elected based orientations, anhree represenubjected to theonformation onergies betweend compared wI).
c features buraints that expnhibitor bindi
Screening andveloped pharm
queries to retrihibitor from aybridge. Chemophoric featuproperties u
rs. Detailed rieved from ea6.
base screening pharmac
x database, 76gh Pharm1, P
mpounds that ophore modelsand, 1 compo
ge database. Fid from the dmolecular doc
r Docking andlar docking it compounds investigate therom the dockion the high GO
nd matching wntative structure energy minimof the hit coen each hit cowith that of th
ut differs in teplain the indung.
d Drug-Like Fmacophore moieve potential two chemicamical compou
ures were retaiusing Lipinski
description ach step of dat
result using thrcophore models
6, 106, and 2Pharm2, and P
are commonls were selecte
ound was chosinally, a total database screcking study.
d Interaction Ecalculations wobtained from
eir binding oriing results, 5 OLD fitness sc
with their pharmres with eachmization to opompounds. Tmpound and rhe original th
erms of inter-uced conform
Filtration odels were utidrug candida
al databases unds mapped ined and verifi’s rule of fion the numtabase screeni
ree receptor-bas
231 compoundPharm3, respely passed throed as hit compsen as hit comof 12 hit comeening proce
Energy were carried m database scientation at thehit compoundcores, proper bmacophore m
h hit compounptimize their b
Then, the interenin were cal
hree inhibitors
-feature mational
lized as ates as a namely well on fied for ive and
mber of ing was
sed
ds were ectively. ough all pounds. mpound
mpounds ess and
out to reening e active ds were binding
mapping. nd were binding eraction lculated s (Table
weresdivtheintselan
Fi
phmaPh
Ph
nointphbothachthe
Inhibitors and Hits
Inh1, Inh2, andInh3 Hit1
Hit2
Hit3
Hit4
Hit5
The interactioere - 57.625spectively. Alversity (Fig. 7e three originteraction enerlected as fina
ntihypertensive
g. 7 Two-dimenon their G
The mappinharmacophore apped over alharm2 and HB
Fig. 8 Pharmacharm2, and c Ph
gr
The binding oot Rep1 wereteraction with
harmacophoricound toward laat is formed
hlorides of Inheir binding mo
TAINTERACTION EN
Rep1
d - 57.62554
- 81.93515
- 78.32083
- 73.46501
- 65.66974
- 58.17558
on energies of54, - 16.326l the 5 hit com
7) were of lownal inhibitorsrgy and reasal hit compoe agent.
nsional chemicGOLD fitness s
ng of this models has re
ll the featuresBA in Pharm3
cophore mappinharm3. HBA, Hreen, magenta, a
orientations oe similar to h the active sc features. In arge hydropho
by interactih1 with the acodes were not
ABLE III NERGIES (KCAL/M
Rep2
4 - 16.3265
5 - 84.6465
- 88.5291
- 75.7259
4 - 69.2833
8 - 66.8589
f Inh1, Inh2, 652, and - 4mpounds of cower interactions, but hit1 sonable bindin
ound and can
al structures of score and bindin
hit compounevealed that ths except for H(Fig. 8).
ng. Hit1 is mapHBD, and HYP and cyan, respe
of the hit1 for each other asite residues Rep1, all 5 h
obic cavity inon of phenyctive site residt consistent w
MOL)
Rep3
52 - 43.927
55 - 93.787
9 - 80.617
94 - 91.957
36 - 74.016
97 - 68.532
and Inh3 with43.92793 kca
onsiderable chn energy valueshowing the ng mode wandidate of po
f the hits filteredng orientation
nd over all his hit compouHBD in Pharm
pped on a Pharmfeatures are sh
ectively
Rep2 and Reand shown to
complementihit compoundn the active sityl moiety witdues. Conseq
with an orienta
793
731
787
759
645
240
h renin al/mol,
hemical es than lowest
as only otential
d based
three und has m1 and
m1, b own in
ep3 but o have ing the ds were te cleft th two
quently, ation of
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PhRm9)
Fisc
hyinthbobenibohibohichbeRanththdimcoen53thpo
PrPrPrFoEdKND
[1]
harm1. This rep1 was optim
modes of hit1 w).
ig. 9 Docking costructures. Bindcompared along
Rep3
Two phenylydrophobic innteraction withhrough oxygenond interactioenzoate moietitrogen and siond interactioit1 and backbond interactioit1 and two hhain oxygen aecause the sulfep3. Althoughnd the active she interaction han - 84.6465ifference in V
mode of the hitonformation nergies of hit3.1862 kcal/mhese results, wotential virtua
This researcrogram (2012rogram (2009-rogram (201oundation of ducation, Scie
Korea. And Next-Generatio
evelopment A
] Murray CJL,disability by Lancet,vol.34
result is becaumized for Inhwere similar to
onformations oding conformatig with hydrogen
are colored in g
l rings of hitnteractions. Thh hydrogen aton atom of mo
ons were founty of hit1 and de chain oxyg
on between hybone oxygen ons between ohydrogen atomatom of Ser2fone moiety wh the numbersite residues ienergy of - 9355 kcal/mol VDW interactt1 that is simioptimized fot1 for Rep2
mol, respectivewe suggest thal lead for reni
ACKNO
ch was suppoR1A1A4A010-0081539), an0-0029084)
f Korea (NRence and Tecthis work
on BioGreen 2Administration
REF
, Lopez AD, “Acause 1990-2020
49, no.9064, pp.14
use the active h1. In Rep2 ano each other an
of hit1 compounions of hit1 in an bond interactigreen and red, r
t1 were mainhe hit1 has forom of backboorpholine mo
nd between oxtwo hydrogen
gen of Ser84. ydrogen atomatom of Gly
oxygen atom oms of backbon230 were onlywas turned the r of hydrogenn Rep2 is mo3.78731 kcal/min Rep2. Thi
tion energies ilar to Inh3 fior Inh3. Theand Rep3 weely. Based onhat hit1 is ofin inhibitor de
OWLEDGMENT orted by Basi013657), Pion
nd Managementhrough the
RF) funded bchnology (ME
was also 21 Program (Pn (RDA) of Re
FERENCES Alternative proje0: Global Burden498-1504, 1997.
site conformand Rep3, the bnd to that of In
nd in two represa Rep2 and b Reng residues. Rerespectively
nly involved rmed hydrogene nitrogen of
oiety. Two hyxygen atom on atoms of baThere is a hy
m of amide moy228. Two hyof sulfone mone nitrogen ay observed inopposite dire
n bonds betweore than that inmol in Rep3 iis is caused because the b
it well the acte VDW inteere - 35.1454n the combinaf novel scaffoesign.
ic Science Rneer Researchnt of Climate C
National Rby the MiniEST) of Repusupported bJ008038) fromepublic of Ko
ections of mortan of Disease Stu
ation of binding
nh3 (Fig.
sentative ep3 are ep2 and
in the en bond f Tyr20 ydrogen of ethyl ackbone ydrogen oiety of ydrogen oiety of nd side
n Rep2, ction in
een hit1 n Rep3, is lower by the
binding tive site eraction 4 and - ation of old and
Research h Center Change
Research stry of ublic of by the m Rural rea.
ality and udy,” The
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