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: chip@bio.gSundarapandian

cience, Gyeongazwa-dong, Ji

under@bio.gnu.acYuno 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: yuno@bio.gnu.of Applied Life

GNU), 501 Jinju-l: minky@bio.gnuon 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

World Academy of Science, Engineering and TechnologyInternational Journal of Medical, Health, Pharmaceutical and Biomedical Engineering Vol:7 No:1, 2013

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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.

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