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Preprint:Pleasenotethatthisarticlehasnotcompletedpeerreview.
ComputationalscreeningapproachesforinvestigatingpotentialactivityofphytoligandsagainstSARS-CoV-2CURRENTSTATUS:POSTED
AcharyaBalkrishnaPatanjaliResearchInstitute
PallaviThakurPatanjaliResearchInstitute
ShivamSinghPatanjaliResearchInstitute
SwamiNarsinghPatanjaliResearchInstitute
NamitaSinghGuruJambheshwarUniversityofScienceandTechnology
RakeshKumarSharmaSaveethaInstituteofMedicalandTechnicalSciences
DOI:10.21203/rs.3.rs-23206/v1
SUBJECTAREASBioinformatics NaturalProductChemistry
KEYWORDSCoronavirus,COVID-19,Drugdesigning,Herbaldrug,Moleculardocking,SARS-CoV-2
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AbstractObjective:SARS-CoV-2causesCOVID-19,alife-threateningrespiratoryillnesswithhighratesof
morbidityandmortality.Asondate,thereisnospecificmedicinetopreventortreatCOVID-19.
Therefore,thereisanacuteneedtoidentifyevidence-basedholisticandsafemitigators.
Methods:Thepresentstudyisaimedtoscreenligandsofherbaloriginusingrationalebased
bioprospectionanalysisandsubsequentlypredicttheirbindingpotentialsubduethemajordrug
targetsfornovelCoronavirusbyemployingcomputer-aidedvirtualscreening.Further,comparative
analysisofthebindingpotentialofanapprovedchemicalanalogueandselectedherballigandswere
alsopredicted.Theselectionofreceptorswasperformedbasedontheirpathophysiologicalrelevance,
asassessedbyaPubMedbasedkeywordhitsmatrixanalysis.ThedruglikelinessandADMETox
descriptorsof24herballigandswerecomputationallypredicted.Dockingstudieswerefurther
conductedwiththosephytoligandsthatqualifiedtheseparameters.Anexistingantimalarialdrug,
hydroxychloroquine,wasalsodockedwithalltheselectedviralreceptorsanditstheoreticalbinding
energywassetupasastandardforcomparisonaswellasscrutinizationofbindingenergiesofthe
phytoligands.
Results:Thedockingstudiessuggestedthattheherballigand,namely,gamma-glutamyl-S-
allylcysteinedemonstratedhighlysignificantbindingenergieswithviralspikeglycoprotein,
endoribonucleaseandmainprotease(bindingenergy≥-490kcal/molforallthetestedviral
receptors).
Conclusion:Gamma-glutamyl-S-allylcysteinedemonstratedmoresignificantbindingpotentialas
comparedtotheknownchemicalanalogue,i.e.,hydroxychloroquine,asobservedinthe
computationaldockingstudies.Thisstudyservestopresentpre-eminentinformationforfurther
clinicalstudieshighlightingtheutilityofherballigandsasprobableleadmoleculesformitigating
novelCoronavirusinfection.
1.IntroductionCoronaviruses(CoVs)representacategoryofinfectiousagents,belongingtothefamily
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Coronaviridae,categorizedintofourgenera,namely,alpha-CoV,beta-CoV,gamma-CoV,anddelta-
CoValongwiththeirsubclasses(Lefkowitzetal.,2018).TheseCoVscontainlargehost-derived
membraneenvelope,helicalnucleocapsid,non-structuralproteins(nsp1–16)andpositivesense,
single-strandedRNA(~26–32kilobases).TheviralRNAiscapableenoughofinitiatinginfectionwithin
thehost,evenwithoutthepresenceofessentialenzymes(Tripp&Tompkins,2018).Sincethe
beginningofthe21stcentury,CoVshavecausedthreezoonoticoutbursts,namely,severeacute
respiratorysyndrome(SARS-CoV;2002–2003),MiddleEastrespiratorysyndrome(MERS-CoV;2012),
andtheCOVID–19,causedbysevereacuterespiratorysyndromeCoronavirus2(SARS-CoV–2;2019)
(Zakietal.,2012;Ouetal.,2020).ThenovelCOVID–19pandemicoriginatedfromtheHubeiprovince
ofCentralChinaduringthelateNovember,2020,withitsepicenterbeinginWuhancityharbouring
nearly11millionpeople(Huetal.,2020).Thisinfectionhasbeengrowingsincethenandhasspread
tomorethan205countries(Cucinotta&Vanelli,2020;WHO,2020).Thetransmissibilityand
penetrancerateofthisinfectionisfrequentlychangingathourlyanddailybasis(Roseretal.,
2020).TheclinicalmanifestationsofthenovelCOVID–19infectionincluderespiratory,gastrointestinal,
nephrologicalandneurologicaletiologies(Balkrishnaetal.,2020).Primarytransmissionofthis
infectioninvolvedzoonoticspreadfromanimals(possiblybats)tohumans(Zhangetal.,2020;Zhou
etal.,2020).Inparticular,theprimarytransmissionofCOVID–19infectionwasapparentlylinkedto
theingestionofliveanimalandseafood(Balkrishnaetal.,2020).AlthoughthebatCoVbears
maximumsimilaritywithSARS-CoV–2,butduetothedifferenceinspecificaminoacidresidues,the
batCoVcouldnotdirectlyinfecthumans,neverthelesstherewouldbeanintermediatehostbetween
batsandhumansforthetransmittanceofthevirus(Cyranoski,2020).Subsequently,secondary
transmissionoccurredamonghumans,leadingtoafasterrateofspreadoftheinfectionwithinthe
community(Guanetal.,2003;Balkrishnaetal.,2020).TheincubationperiodofSARS-CoV–2ranges
between1–14dayswiththemedianincubationperiodofabout5days.Aninfectedpersonwill
apparentlyshowsymptomsafterthemedianincubationperiod.However,theinfectedindividualcan
spreadtheinfectionsincecontractingthedisease.TheprimarymodeoftransmissionofCOVID–19
infectionisthroughcontactwiththeinfectedindividualsand/orbyrespiratorydroplets(Carlosetal.,
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2020).SomestudieshavealsoproposedthatSARS-CoV–2mayalsopropagatebythefaecal
shreddingofinfectedorevencuredindividuals(Yeoetal.,2020).
TheCOVID–19infectionhasbecomeaglobalhealthchallengeowingtoitshighmorbidity(~3.17%);
transmissionrate(~2.2perpatient)(Sunetal.,2020);longerhalf-lifeofvirus(~4–72hours);and
asymptomaticmodeoftransmission(~2–14daysofincubation)(Huangetal.,2020;Balkrishnaetal.,
2020).ThemostcommonindicationsofCOVID–19compriseoffever,nasopharyngealcongestion,dry
cough,sorethroat,dyspnoea,diarrhoea,neurologicaldisordersandmultipleorganfailure.However,
somepatientsactasasymptomaticcarriersofthediseasewhicharemostdifficulttodiagnoseand
consequentlytreat.Eventhoughthedistinctsymptomsseemtobemild,butcanrapidlyenhanceto
severeconditions,andevendeath(Huangetal.,2020;Kim,2020).
TheexactmolecularpathogenesisofSARS-CoV–2isyetnotknownwithcertainty;however,ithas
beenproposedthatviralpathogenesisistriggeredbythereleaseofproinflammatorycytokinesthat
areassociatedwiththeactivationofseveralsignalingpathways,namely,TLRs-dependentIFN
inductionpathways(IRF–3/7&NF-Kβ)andmyeloid-differentiationprimaryresponse88pathways(ATF–
2&AP–1).TheprimaryvirulencefactorsofthevirusincludeSARS-CoV–2spikeglycoprotein(viral
envelopeproteinresponsibleforviralattachmentandentrywithinthehostcells);viralnuclease
(NSP15endoribonucleaseresponsibleformediatingviralcapsidformation);andprotease(Main
Protease3CLproresponsibleforviralcapsidformation)(Chenetal.,2020).SARS-CoV–2mainly
targetsthealveolarandbronchialepithelialcells.TheviralspikeglycoproteininteractswiththeACE–
2receptorsofthehostcell,therebymediatingtheviralentry.Afterwards,thereoccursanelevated
releaseofproinflammatorycytokines(IL–6andIL–12),chemokines(IL–8,CCL–2andCXCL10)and
interferon.Thevirusgetsholdofthehostcellmachineryandmanipulatesitfordrivingtheprocessof
viralreplication.Virusalsoattemptstoinhibittheproductionofinterferonandproinflammatory
cytokinesbymeansofRNAhelicasesandnon-structuralproteins(Nsp1,3,7&15).Subsequently,
theviralloadkeepsonrisingwithinthehostsystem,ultimatelyleadingtoviraemia(Figure1)..
Meanwhile,thehostimmunesystemalsostrivestofightback.Thefurtherspreadofviruswithinthe
hostsystemdependsontheimmunologicalstatusofthehost.Immunomodulatoryremediesplaya
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pivotalroleatthisphase,whereinsuchmoietiesmaymediatethetransformationofan
immunocompromisedindividualtoanimmunocompetentone.
Although,thestructureandsequenceofSARS-CoV–2havebeenidentifiedanddrugscreening
followedbyclinicaltrialsarecontinuouslybeingconductedbytargetingthesevirulencefactors.
However,therearenoapproveddrugsforeffectivelymanagingCOVID–19infection,probablydueto
unidentifieddynamicpathophysiology;highmutagenicityofthevirus;andadversesideeffectsof
earlierknownCoronavirusvaccinesanddrugs(Guoetal.,2020;Senathilakeetal.,2020;Wangetal.,
2020).Inlightoftheoutbreak,severaltreatmentregimeshavebeenproposed,includingtraditional
medicinalmoieties,thathavebeenextensivelyusedduringtheformerepidemicoutbreaks,suchas
SARSandH1N1influenza(Zakaryanetal.,2017;Guerrieroetal.,2018;Yangetal.,2018;Angetal.,
2020;Joetal.,2020).Herbalsprovideauniquesolutionintermsoftheirnegligiblesideeffects,
synergisticactivity,broadspectrumtherapeuticabilityandimmunomodulationeffects(Thakuretal.,
2016).
Inthepresentstudy,thedrug-likepotentialofsecondarymetabolitesofherbaloriginwillbestudied
bytargetingSARS-CoV–2spikeglycoprotein(S2),viralnuclease(NSP15endoribonuclease)and
protease(MainProtease3CLpro).Theselectionofherbalsecondarymetabolitesisbasedon
investigatingthepotencyofvariousherbalsshowingresistancetowardsviralvirulencefactorsas
indicatedbyvariousscientificsearchenginesbasedonpriorityindexing.Alltheselectedherbal
moietiesweresubjectedtomoleculardockingusingHexsoftware,soastoassesstheinteractionsof
phytoligandswiththeviralreceptors.Furtherscreeningofthepropitiousherballeadswasconducted
byassessingtheinsilicotoxicityandLipinskiscorebaseddruglikeliness.Bulkoutliersshowinghigh
toxicity/mutagenicityorviolatingtheLipinskiruleswereeliminated.Subsequently,thedrug-receptor
interactionofthefilteredherbalmoietieswasstudiedsoastoobtainaleadmoleculewhichcouldbe
furthertestedatpreclinicalandclinicallevels.Althoughsearchforpotentialleadstargetingthenovel
Coronaviruswillcontinueperpetually,buttheseherballeadsmayservetobehighlybeneficialowing
totheirantiviralactivities,potentiatingnatureandsymptomaticreliefprovisioncapabilities,
presentedalongwithlimitedtoxicitiesandcomprehensivetreatmentstrategy.
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2.MaterialsAndMethods2.1Selectionandpreparationofviralvirulencefactorsasreceptors
ThereceptorswereselectedbasedontheirroleincausingviralinfectionbySARS-CoV–2virus.These
receptorswereassignedaweightagescoreonthebasisoftheirpathophysiologicalrelevance.Such
pertinencewasassessedbyconductingakeywordhitsscoringmatrixusingPubMedsearchengine.
Therandomkeywordsearchwasconductedusingcombinationkeywordas‘virulencefactorinhibition
+antiviralactivity’.Thefirst20hitsprovidedbythesearchengine,workingontheprincipleof
priorityindexing,werebasedonthenumberoftimesawebsiteisclicked(Qinetal.,2005).This
samplesetbasedanalysiswasusedtoevaluatethenetweightagelinkedtoeachvirulencefactor,
usingthefollowingformula:
Thecrystalstructuresoftheselectedrelevantproteintargets,namely,SARS-CoV–2spike
glycoprotein(S2;PDBcode:6VSB),viralnuclease(NSP15endoribonuclease;PDBcode:6VWW)and
protease(MainProtease3CLpro;PDBcode:1Q2W)wereobtainedfromRCSBProteinDataBank
(https://www.rcsb.org/).ThesestructureswereexaminedcriticallyusingRamachandranPlotby
ProChecksoastovalidatethemodeledproteinstructuresbasedontheφ(phi),ψ(psi)andω(omega)
angles,therebyinspectingthequalityofthetargetproteinstructuresselectedfordockingstudies.
Furthermore,hydrogenatomswereintroducedinallthese3DstructuresusingArgusLab(4.0.1),so
astocustomizetheviralreceptorsforrigiddocking
(http://www.arguslab.com/arguslab.com/ArgusLab.html).
2.2Activesiteanalysisofviralvirulencefactors
ThepredictionofactivesitesofviralreceptorswasaccomplishedbyDoGSiteScorer,andthe
Cartesiancoordinatesx,y,z(activesites)foreffectivedockingwerevisualizedinArgusLab.These
regionswerefurtherusedforthegenerationofgridboxfordockingstudiesbyHexCuda8.0.0.
PredictionswithDoGSiteScorerwerebasedonthedifferenceofGaussianfiltertodetectpotential
pocketsontheproteinsurfacesandtherebysplittingthemintovarioussub-pockets.Subsequently,
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globalproperties,describingthesize,shapeandchemicalfeaturesofthepredictedpocketswere
calculatedsoastoestimatesimplescoreforeachpocket,basedonalinearcombinationofthree
descriptors,i.e.,,volume,hydrophobicityandenclosure.Foreachqueriedinputstructure,a
druggabilityscorebetween0-to–1wasobtained.Higherthedruggabilityscore,higherthe
physiologicalrelevanceofthepocketaspotentialtarget.Theactivesiteswithbestdruggability
scoreswereenvisagedasanessentialrequirementforthecreationofgridinselectedviralreceptors
(Volkameretal.,2012).
2.3Selectionandpreparationofherbalsaspromisinganti-SARS-CoV–2candidates
Atotalof24bioactivecompoundsfromnaturallyavailablemedicinalplantswereselectedby
employingabiostatisticalmatrixbasedanalysis.Basedontheunderstandingofthe
pathophysiologicaltargetsofthenovelCoronavirus,herbalcandidatesexhibitinginhibitoryproperties
specifictotheviralvirulencefactorsweresearchedinthePubMedrepository.Thedescriptorsused
forconductingPubMedsearchwiththehelpofextensiveliteraturesurgeincludedkeywordsas
‘virulencefactorinhibition+herbalmoiety’.Binarycoefficientfortheherbalmoietieswerecalculated
byassessingthepresenceorabsenceofparticularinhibitingpropertyexhibitedagainstindividual
physiologicaltarget.Thebinaryscoreforeachplantrangedfrom0to6,whereinthemediancut-off
valuewasselectedas3.Plantshavingabinaryscore≥3wereconsideredforfurtherweightage
basedmatrixanalysis,whereinthebinaryscoreofeachplantwasmultipliedwiththerelevancescore
oftheviralvirulencefactor(RefertoSection2.1).Ultimately,afuzzysetmembershipanalysiswas
conductedinordertoobtainauniversalscoreforeachplant.Thefuzzysetscorerangedbetween0
and1,whereintheplantswithafuzzyscoregreaterthan0.5werefurtherselectedforassessingtheir
specificanti-SARS-CoV–2activity.
where,μSrepresentsthedesirabilityvaluesofmembersofthefuzzysetS;min(S)andmax(S)are
minimumandmaximumvalues,respectively,inthefuzzysetS.
Predominantphytochemicaloftheselectedplants(withfuzzyscore≥0.5)wereidentifiedby
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extensiveliteraturesurge.Thethree-dimensionalstructuresofallthesebioactivemoleculesaswell
asthereferencedrugcompound(hydroxychloroquinewhichisunderconsiderationasaremedyfor
novelCOVID–19viraldisease)wereretrievedfromPubChemdatabase.Hydrogenatomswere
introducedintotheligandsstructureusingArguslab(4.0.1),soastocustomizethemforrigid
docking.ThehydrogenatedligandmoleculeswerethenconvertedintopdbformatusingOpenBabel
(2.4)interface(openbabel.org/docs/dev/OpenBabel.pdf),asrequiredforrigiddocking.Similarly,3D
structureofstandardchemotherapeuticagent(hydroxychloroquine)wasalsocustomizedfordocking
(Modietal.,2013;Chakotiyaetal.,2014).
2.4Insilicopharmacokineticanalysis
2.4.1DrugLikeliness
Druglikelinessoftheselectedphytoligands(~17compounds)wasassessedbyusingDruglikeness
toolDruLiTo,whichisanopensourcevirtualscreeningtoolforcalculatingLipinski’sruleoffive,i.e.,
molecularweight,numberofhydrogenbonddonors,numberofhydrogenbondacceptorsandLogP
value.Violationofmorethanonerulewouldcauseexclusionofthesaidphytochemical.Restofthe
selectedphytoligandsweresubjectedtoADMEToxanalysis(Kriszhnamurthietal.,2018)
2.4.2ADMEToxAnalysis
TheADMETox(Absorption,Distribution,Metabolism,ExcretionandToxicity)descriptorsofthe
selectedphytocompoundswerepredictedbyconductingadmetSAR.Theestimationoftheprobability
valuesofthecompoundsfordiverseprofilesincludinghumanoralbioavailability,humanepithelial
colorectaladenocarcinomacell(CaCo2)permeability,logPforsubstratesandinhibitors,predicted
aqueoussolubilityanddifferenttoxicityprofilesintermsofAMEStoxicityandoraltoxicity(LC50and
LD50values)werecomputationallypredicted.Phytomoleculesexhibitingtoxicprofileasassessedin
theinsilicotoxicityanalyseswereexcludedfromthestudy.Restoftheselectedphytoligandswere
subjectedtomoleculardockinganalysis(Nishaetal.,2016).
2.5MolecularDockingandLigandReceptorBindinganalysis
Thedockinganalysisofpdbstructuresofselectedphytoligands(excludingtheLipinskiruleand
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ADMEToxviolatingmoieties)withviralreceptors(spikeglycoprotein,viralnucleaseandviralmain
protease)wascarriedbyHexCuda8.0.0software.ReceptorandLigandfileswereimportedinthe
software.ThegriddimensionofdockingwasdefinedaccordingthetothebindingsiteanalysisofDoG
SiteScorer.GraphicsettingsandDockingparameterswerecustomizedsoastocalculatethebinding
energies(Evalues)ofligandreceptordocking.Theparametersusedforthedockingprocesswereset
as(i)Correlationtype:Shape+Electro+DARS,(ii)FFTmode:3Dfastlite,(iii)GridDimension:0.7,
(iv)Receptorrange:180°,(v)Ligandrange:180°,(vi)Twistrange:360°.Thebestdocked
conformationswithlowestdockingenergywereselectedforfurtherMDsimulationsusingPoseView
forcreatingposedepictionsofselectedligand-receptorbinding(Harikaetal.,2017).
3.ResultsAndDiscussion3.1Selectionofviralvirulencefactors
Onthebasisofthekeywordhitsscoringresults,weightagewasgiventovariousparametersselected
forscreeningofherballeadswithrespecttoantiviralactivity.Weightagewasdecidedaccordingto
thepercentagerelevanceobtainedforeachparameter,asgiveninTable1.Highestpercentage
relevancewasobtainedforviralspikeglycoproteininhibitor,followedbyotherparameterslikeviral
endonucleaseinhibitorandviralproteaseinhibitor.Dockingstudieswerehenceperformedwiththese
viralreceptors,namely,SARS-CoV–2spikeglycoprotein(S2),viralnuclease(NSP15endoribonuclease)
andprotease(MainProtease3CLpro).
3.2Qualityassessmentofviralvirulencefactors
Thefirstqualityassessmentoftheselectedviralvirulencefactors(SARS-CoV–2spikeglycoproteinS2,
viralNSP15endoribonucleaseandmainprotease3CLpro)wascarriedoutusingRamachandranplot
analysiscomputedwithProCheck.TheanalysisshowedthatresiduesofSARS-CoV–2spike
glycoproteinS2,viralNSP15endoribonucleaseandmainprotease3CLprointhemostfavorable
regionwere84%,93.1%and89.6%,respectively.Moreover,intheadditionallyallowedregions,
nearly15.9%,6.9%and9.6%residuesofSARS-CoV–2spikeglycoproteinS2,viralNSP15
endoribonucleaseandmainprotease3CLprowerefound,respectively.ThetotalqualityG-factorof
themodeledviralreceptorssuggestedthatallofthemexhibitedfavourablestereo-chemical
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parametersandhence,the3Dstructuresofalltheseviralreceptorscorrespondtohigh-probability
conformationformoleculardocking.TheProCheckstereo-chemicalanalysisalsoshowednobad
contactsandscoresformain-chainorside-chainparameters.Thedetailedsecondarystructural
investigationoftheSARS-CoV–2spikeglycoproteinS2withPDBsumserverrevealedthat313(10.77%)
residueswereinstrands,2112(72.70%)residueswereinα-helices,78(2.68%)residueswereinβ-
turnsand402(13.83%)residueswereinotherconformations.Similarly,PDBsumsecondarystructure
ofNSP15endoribonucleaserevealedthat72(10.34%)residueswereinstrands,577(82.90%)
residueswereinα-helices,4(0.57%)residueswereinβ-turnsand43(6.17%)residueswereinother
conformations.Moreover,PDBsumsecondarystructureofmainprotease3CLprorevealedthat75
(12.66%)residueswereinstrands,457(77.19%)residueswereinα-helices,7(1.18%)residueswere
inβ-turnsand53(8.95%)residueswereinotherconformations(Figure2)..Thus,theaboveanalysis
suggeststhatthebackboneconformationandnon-bondedinteractionsofalltheselectedviral
receptorswerefoundtobewithinreasonablerangeandhencecanbeprocessedforfurtherdocking
analysis(Kumaretal.,2011).
3.3Activesiteanalysisofviralreceptors
ActivesiteanalysisofSARS-CoV–2spikeglycoprotein(S2),viralnuclease(NSP15endoribonuclease)
andprotease(MainProtease3CLpro)asconductedbyDoGSiteScorerindicatedthatthereare
variousactivepocketswithinthestudiedviralvirulencefactorswithdruggabilityrangingfrom0.12to
0.86(Table2)..ItwasfoundthatpocketsP_11(Drugscore:0.847),P_1(Drugscore:0.860)andP_0
(Drugscore:0.805)wereenergeticallyfavourableforperformingfurthermoleculardockingstudies
withtheviralreceptorsbeingspikeglycoprotein,NSP15endoribonucleaseandMainProtease3CLpro,
respectively.Whileconductingtheactivesiteanalysis,theDoGSiteScorertoolanalysedtheheavy
atomcoordinatesonthesurfaceofthe3Dstructureoftherespectiveviralreceptors.Dependingon
theseatomiccoordinates,ahypotheticalgridwasspannedbyoutrulingthechancesofanyspatial
overlapofthegridwiththeheavyatoms.Furthermore,thetoolengagesinapplyingaGaussianfilter
tothedefinedgrids,soastoidentifysphericalpocketsofbinding.Druggabilityscore(0–1)ofthe
selectedsphericalpocketsarededucedonthebasisoftheirsurfacearea,volume,enclosureand
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hydrophobicity.Asageneralrule,higherdruggabilityscoreisindicativeofamoredruggablepocket
(Volkameretal.,2012).
3.4Selectionofherbalsaspromisinganti-SARS-CoV–2candidates
Extensiveliteraturesurgecombinedwithamatrixbasedanalysiswasconductedforselectionof
plantshavingprobableutilityagainstSARS-CoV–2.Theparametersforselectingtheherbalsincluded-
a)ethnopharmacologicalimportanceoftheplant;b)priorpharmaco-therapeuticinvestigationsofthe
plant;andc)symptomaticreliefprovidingcapabilitiesoftheplant.Binary,weightageandfuzzyscore
analyseswereconductedforalltheplantssoastoscreenoutherbalsexhibitingprobableanti-SARS-
CoV–2activity.Plantsshowingpositiveassessmentformorethan03parameters,reportedinPubMed
searchengine(n=first20hits)wereselectedforfurtherinsilicoanalysis.Therationaleforselected
plantsalongwiththeirbinary,weightageandfuzzyscoreshasbeenexplainedinTable3.
Predominantphytocompoundsoftheselectedherbswerealsoidentifiedbyliteraturesurgeandthe
respectivechemicalstructureswereretrievedfromPubChemdatabase(Chakotiyaetal.,2014).
3.5Pharmacokineticdescriptorsofphytoligands
Druglikelinesscharacteristicsofthebioactivephytoligandswereassessedbyemployingastep-wise
filteringstrategy,whereinvariousphysiochemicalpropertiessuchaslogP,H-bondacceptor,H-bond
donor,molecularweight,acidicgroups,aromaticrings,numberofrotatablebondsandchains,
numberofhydrogenbondsandmolarrefractivitieswerepredictedsoastoevaluatethedruglike
behaviorofthephytoligand.AccordingtotheLipinski’srule,adruglikemoietyshouldhavealow
molecularweight(≤500D),logPvalue≤5,numberofhydrogenbondacceptors≤10,andnumberof
hydrogenbonddonors≤5.Abioactivedruggablemoleculeshouldensuetoatleast4ofthe5Lipinski
rules(Zhang&Wilkinson,2007).TheLipinskiscoresfortheselectedphytomoleculeswerefoundto
bewithinacceptablerangesaselucidatedinTable4.
3.6ADMEToxpredictionofphytoligands
TheresultsofadmetSARpredictionshowingtheprobabilityvaluesaresummarizedinTable5.The
phytoligandsviolatinganyoftheADMEToxdescriptors(amentoflavone,butenolide,malicacid,β-
myrcene,paeoniflorin)wereexcludedatthisstepitself.Restofthephytoligands(~12moieties)did
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notexhibitanymutagenicortoxicprofile.Basedonthepredictedprobabilityvalues,theselected
phytoligandswereknowntogetabsorbedefficientlybytheintestinalepitheliumasthevaluesfor
CaCo2permeabilityandintestinalabsorptionwerefoundtobewithinpermissiblerange(~absorption
value≥0.5).Suchpredictedabsorptionvaluessuggestthattheselectedphytoligandsmaygeteasily
transportedaftergettingabsorbedinthehumanbody.Theaqueoussolubilityoftheselected
phytoligandswasalsopredictedtobeacceptable(logS≥-4).Moreover,alltheselectedphytoligands
alsoexhibitedefficientbindingwiththeplasmaprotein,therebyensuringefficientdistributionofthe
probabledrugmoieties(Plasmaproteinbindingvalue≥0.5).Asfarasmetabolismisconcerned,
inhibitionpredictionsforcytochromeP450(CYP3A4)superfamilywereconducted.Noneofthe
selectedphytomoleculesexhibitedanyinhibitionofCYP3A4,thusensuringproperdrugmetabolism
andnoexcessivedrugaccumulationaswellastoxicity.MostoftheselecteddrugsalsoexhibitedP-
glycoproteinmodulationactivity(~P-glycoproteinactivationvalue≥0.5),therebymediatingactive
transportandmetabolismofthedrugmoieties.Othertoxicityandexcretioncharacteristicsalso
suggestedthattheselectedphytoligandsdidnotexhibitanysignsoforaltoxicity(~acuteoral
toxicityvalue≥1.0Kg/mol)andmutagenicity(Amesmutagenicityvalue≥0.5),thusmakingthe
respectivephytoligandsasprobableleadmolecules(Nishaetal.,2016).
3.7MolecularDockinganalysis
Dockingresultsoftheviralvirulencefactors,namely,spikeglycoprotein,NSP15endoribonuclease
andMainProtease3CLpro;andtheselectedphytoligands(~12phytomolecules)areshowninTable
6.ThesedockingbasedEvalueshavealsobeencomparedwiththatofthestandarddrug,i.e.,
hydroxychloroquine.HexbaseddockingresultsrevealedthattheE-valueofdockingofgamma-
glutamyl-S-allylcysteineandsalvianolicacidwithalltheselectedviralreceptors(viralmainprotease
3CLpro,spikeglycoproteinandNSP15endoribonuclease)wassignificantlybetterascomparedto
hydroxychloroquine.Severalotherphytoligandsalsoshowedcomparablebindingenergieswith
respecttoatleastoneoftheviralreceptors;however,noneoftheotherphyto-moietiesexhibited
holisticdockingabilities.Hence,itisobviousfromtheE-valuesthatgamma-glutamyl-S-allylcysteine
andsalvianolicacidbindspontaneouslyandirreversiblytoallthetestedviralreceptors,thereby
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blockingthespreadandreplicationofthenovelCOVID–19virus.Moreover,thebindingefficiencyof
gamma-glutamyl-S-allylcysteinehasbeenfoundtobeexceedinglybetterthanthatofsalvianolicacid.
Suchsignificantbindingaffinityofgamma-glutamyl-S-allylcysteineandsalvianolicacidpresumably
indicatestheprobablemechanismofactionofthesephytomoleculesasviralprotease,spike
glycoproteinandendoribonucleaseinhibitors.Suchefficientbindingenergyoftheseherbalmoieties
indicatesthemtobecomparablewiththatofhydroxychloroquinewhichhasbeenproposedasthe
cornerstoneforCOVID–19therapy.
Viralspikeglycoproteinisrequiredforinitiatingtheattachmentandentryofvirusintothehostcell.
Moreover,viralproteaseisfundamentalforcontinuingthevirallifecycleofSARS-CoV–2asitis
requiredbythevirustocatalyzethecleavageofviralpolyproteinprecursorswhichareultimately
necessaryforviralcapsidformationandenzymeproduction(Anandetal.,2003).Similarly,viral
endonucleasesarenecessaryforcatalyzingtheprocessingofviralRNAsandhencearerequiredfor
enduringtheprocessofviralreplication(Balkrishnaetal.,2020).Henceforth,gamma-glutamyl-S-
allylcysteineandsalvianolicacidcontingentlyinhibitstheentryofSARS-CoV–2andalsoinactivates
theviralproteaseaswellasendoribonuclease,therebyinhibitingtheprocessofviralcapsidformation
andreplicationaltogether.Earlierstudieshavealsoindicatedthatgammaglutamylcysteineester
derivativescouldinhibittheHIV–1genetranscription,wherein,itprobablyrestrainedtheoxygenfree
radical-mediatedactivationofthenuclearfactor-kappaB(NF-κB).Moreover,thismoietyhasalonger
plasmalife,bioavailabilityandmembranepermeability,therebymakingitasuitabledrugcandidate
withsustainedantiviralaction(Kubotaetal.,1998).
3.8PhytoligandandViralreceptorbindingposedepictions
Thebestdockedconformationswithlowestdockingenergy,i.e.,gamma-glutamyl-S-allylcysteineand
salvianolicacidwereselectedforfurtherMDsimulationsusingPoseViewforcreatingposedepictions
ofselectedligand-receptorbinding.Uponassessingthebindingposeandelectrostaticbridging
interactions,itwasfoundthatonlygamma-glutamyl-S-allylcysteine(Herbalsource:Alliumsativum)
wasabletofitintotheactivebindingpocketsoftheviralreceptors,whereas,salvianolicacidcould
notestablishanirreversibleandspontaneousbondwiththeviralreceptors.Theorientationalbinding
14
ofgamma-glutamyl-S-allylcysteineandtheviralreceptorsshowingtheposeviewandresidue
interactionshavebeendepictedinFigure3.Itwasobservedthattheamidegroupofgamma-
glutamyl-S-allylcysteineformedahydrogenbondwiththeamideresidueofglutamineaminoacid
(1071stposition)foundintheviralspikeglycoprotein.Chemicalbridgingofgamma-glutamyl-S-
allylcysteineandglutamicacidresiduesofviralendoribonucleasepresentasimilarcasewhere
glutamicacidresidues(45thposition)wereinvariablyboundandneutralized,therebypossibly
neutralizingtheCOVID–19virus.Inearlierstudiesithasbeenfoundthatmutationoranychangein
theglutamineorglutamicacidresiduesofEbolavirusspikeglycoproteincausesviralneutralization
(Reynard&Volchkov,2015).Similarly,thehydroxylgroupofgamma-glutamyl-S-allylcysteineformed
ahydrogenbondwithcarbonylgroupofprolineaminoacid(108thposition)ofviralmainprotease.
Previousstudieshavealsosuggestedthattheprolineaminoacidresiduesfoundintheconserved
domainsofHIVviralinfectivityfactor(Vif)aretherapeutictargetsforneutralizingthehuman
immunodeficiencyvirus(Ralphetal.,2020).Inconclusion,thebindinginteractionsofgamma-
glutamyl-S-allylcysteinewithviralmainprotease,spikeglycoproteinandendoribonucleasesuggestit
tobeapromisingdrugcandidateformitigatingnovelCoronavirusinfection.
4.ConclusionThenovelCoronavirusinfectionaccountsforinnumerabledeathsworldwide,andthereisyetno
approvedprophylacticvaccineorspecifictreatmentavailable.Basedonevidencefromlaboratory,
animalandclinicalstudies,Hydroxychloroquineisoneofthetreatmentoptionsselectedin
‘Solidarity’-aninternationalclinicaltrialtohelpfindaneffectivetreatmentforCOVID–19,launchedby
theWorldHealthOrganizationandpartners.However,therepurposeddrug,hydroxychloroquinemay
causeadversedrugreactionsandcontraindicationsincludingcardiomyopathy,fulminanthepatic
failure,vertigoandotherallergicreactions.Undersuchcircumstances,thereisanurgentneedfor
screeningnovelnaturalleadsthatexhibitspecificantiviralactivitiesagainstSARS-CoV–2.Thepresent
studysuggestedthatphytoligandsderivedfrommedicinalherbsexhibitedpotentialbinding
propertiestowardmajorSARS-CoV–2virulencefactors.Selectedphytomoleculeswerefurther
screenedonthebasisofacceptablepharmacokineticanddrug-likeproperties,therebymakingthem
15
safelyexploitableforCoronavirusmitigationsystem.Thecurrentstudyshowedthatgamma-glutamyl-
S-allylcysteine(GGA)specificallyexhibitedthemostsignificantbindingenergyanddockingpose
towardthemajorviralvirulencefactors(EvalueGGA+spikeglycoprotein=–578.57Kcal/mol;EvalueGGA
+viralmainprotease=–493.53Kcal/mol;EvalueGGA+endoribonuclease=–825.00Kcal/mol)incomparison
withtheknownchemicalmoietyhydroxychloroquine(EvalueHCQ+spikeglycoprotein=–207.47Kcal/mol;
EvalueHCQ+viralmainprotease=–235.48Kcal/mol;EvalueHCQ+endoribonuclease=–213.54Kcal/mol).
Hence,thecurrentstudyprovidesimplicationtowardthepossibleusageofgamma-glutamyl-S-
allylcysteine(Herbalsource:Alliumsativum)asanovelandprospectivedrugcandidate.This
phytomoleculeisalsofoundinotherspeciesoftheAlliumgenus(A.cepaandA.schoenoprasum)..In
viewofthecurrentviralpandemicanddearthofeffectivetherapy,furtherstudiesshouldbeurgently
undertakensoastoexplorethetherapeuticpotentialofgamma-glutamyl-S-allylcysteineagainst
SARS-CoV–2.
AbbreviationsACE2:Angiotensinconvertingenzyme2;ADMETox:Adsorption,Distribution,Metabolism,Toxicity;AP–
1:Activatorprotein1;ATF–2:Activatingtranscriptionfactor2;CoV:Coronavirus;COVID:Coronavirus
disease;DARS:DecoysasReferenceState;DoG:DifferenceofGaussian;FFT:FastFourierTransform;
GGA:Gamma-glutamyl-S-allylcysteine;GPCR:G-proteincoupledreceptor;HCQ:Hydroxychloroquine;
IRF:Interferonregulatoryfactor;MERS:MiddleEastRespiratorySyndrome;NF-Kβ:NuclearFactor
kappa-light-chain(β);NSP:Nonstructuralprotein;O.E.C.D.:OrganisationforEconomicCo-operation
andDevelopment;PDB:ProteinDataBank;QSAR:QuantitativeStructureActivityRelationship;RCSB:
RoyalCollaborativeStructuralBiology;SARS:SevereAcuteRespiratorySyndrome;TPSA:Totalpolar
surfacearea;VIF:Viralinfectivityfactor
DeclarationsAuthorContributions
ABandRKSconceivedthepresentedresearch.PT,SSandSNDanalyzedtheinformation,generated
theartwork,andco-wrotethemanuscript.AB,NSandRKSinvestigatedandsupervisedthefindingsof
thework.RKSprovidedcriticalrevisionofthisarticle,andapprovedthemanuscriptforsubmission.
16
Allauthorsagreedwiththefinalversionofthismanuscript.
Funding
Noexternalfundinghasbeenreceived.
EthicsStatement
Theauthorsconfirmthattheethicalpoliciesofthejournal,asnotedonthejournal’sauthorguidelines
page,havebeenadheredto.Noethicalapprovalwasrequiredasthisisaninsilicostudy.
ConflictofInterest
Theauthorsdeclarethattheresearchwasconductedintheabsenceofanycommercialorfinancial
relationshipsthatcouldbeconstruedasapotentialconflictofinterest.
Acknowledgments
TheauthorsaregratefultoSwamiRamdevJiforinstitutionalresearchfacilitiesandsupports.Authors
gratefullyacknowledgetheeffortsofcolleaguesofPatanjaliResearchInstitutefortheirhelpindata
collectionandprocessing.WearealsothankfultoMr.GaganKumarandMr.LalitMohanfortheirswift
administrativesupportsandencouragements.
DataAvailabilityStatement
Alldatageneratedoranalysedduringthisstudyareincludedinthispublishedarticle.
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TablesTable 1. Selection of COVID-19 virulence factors on the basis of relevance score as assessed bykeywordhitsscoringmatrix
21
Parameter Rationaleofselection TotalNo.ofHits(N)
HitsScreened(n)
RelevantHits(r)
PercentageRelevance†
ViralSpikeGlycoproteininhibitor
Enveloped viruses enter cells by viralglycoprotein-mediated binding (Viral spikes –S proteins) to host cells and subsequentfusionof virusandhost cellmembranes (Liuetal.,2020).
20 20 18 90
ViralEndoribonucleaseinhibitor
Endoribonuclease catalyses the processingand degradation of both cellular and viralRNAs, thus determining the amount andfunctionality of specific RNA molecules in acell at any given time. It degrades the hostmRNA,whilecleavestheprecursorviralRNAstoproduceactivegeneticmaterial(Balkrishnaetal.,2020).
103 20 13 65
ViralProteaseinhibitor
Viral proteasesareenzymesencodedby thegenetic material of viral pathogens so as tocatalyse the cleavage of specific peptidebonds in viral polyprotein precursors or incellular proteins. The protease cleaves theprecursor viral polyprotein to producefunctional proteins and enzymes (Jo et al.,2020).
1516 20 12 60
Anti-bronchitisHerb
Coronavirus causes respiratory illnesses.Hence, herbs providing symptomatic reliefagainstrespiratorysymptomsmightprovetobebeneficial(Carlosetal.,2020).
551 20 9 45
Anti-gastroenteritisHerb
Coronavirusalsocausesgastroenteritis.Thus,herbs providing symptomatic relief againstgastrointestinal ailments might prove to bebeneficial(Chenetal.,2020).
52 20 8 40
Interferonregulatoryherb
Interferons (IFNs) are cytokines which areused for communication between cells totriggertheprotectivedefenseoftheimmunesystem that help eradicate pathogens,Coronavirus in this case (Balkrishna et al.,2020).
952 20 6 30
Duetotechnicallimitations,table2isonlyavailableasadownloadinthesupplemental
22
filessection.
Table3.SelectedHerbalmoietiesshowingprobableantiviralutilityasassessedbyemployingextensiveliteraturesurge.
PlantSource§ Predominantphytocompound
Activityexplored† BinaryScore
WeightageScore
FuzzyScore*
ProbableantiviralutilityS2 NSP
153CLpro
AlismacanaliculatumA.Braun&C.D.Bouché
AlisolA24-Acetate + − − 2 1.16 0.46 Anti-influenzaactivityobserved as theherbal moietyinactivates thehemagglutininspikereceptor.
AlliumcepaL. Allicin + + + 3 1.66 0.66 Hinders virusattachment tohost cell, altertranscriptionandtranslation ofviral genome inhost cell andalso affect viralassembly.
AlliumsativumL. Gamma-Glutamyl-S-allylcysteine
+ + + 6 2.49 1 Actsasproteaseinhibitormainly.
ArtemisiacapillarisThunb.
Beta-caryophyllene + − + 5 1.91 0.76 Symptomaticalleviation incaseofhepatitisvirusinfection.
ArtemisiacaruifoliaBuch.-Ham.exRoxb.
Caruilignan + − + 3 1.25 0.5 Anti-influenzaand anti-herpessimplex virusactivity; alsoinhibits HIV-1protease.
AsparagusracemosusWilld.
Isoasparagine − − − 3 0.40 0.3 Symptomaticalleviation incase of herpesvirusinfection.
BerberisaristataDC. Berberine + − + 5 1.91 0.76 Inhibitsenterovirus 71entry andreplication bydownregulatingthe MEK/ERKsignalingpathway andautophagy.
BoswelliaserrataRoxb. 11-keto-beta-boswellicacid
+ − + 3 1.66 0.66 InhibitsChikungunyaand Vesicularstomatitis virusinfections byblocking theirentry.
Camelliasinensis(L.) Quercetin + − + 5 1.91 0.76 Suppressed
23
Camelliasinensis(L.)Kuntze
Quercetin + − + 5 1.91 0.76 SuppressedHepatitisCvirusentry, and alsoinhibited viralRNAreplication.
ChlorophytumborivilianumSantapau&R.R.Fern.
Neotigogenin − − − 2 0.40 0.3 Cytokinemodulatingpotential.
CurcumalongaL. Curcumin + − + 5 1.91 0.76 Inhibits entry ofChikungunyaand Vesicularstomatitisvirus.
EpimediumflavumStearn
Wushanicariin + − − 1 1 0.4 Induced thesecretionoftypeI IFN and pro-inflammatorycytokines.
GingkobilobaL. Amentoflavone + − + 4 1.66 0.66 Inhibits viralprotease,specifically incase of HIVinfection.
HouttuyniacordataThunb.
β-myrcene + − + 5 1.91 0.76 Inactivation of3C-likeproteinase ofmurineCoronavirus anddenguevirus.
MelissaofficinalisL. Citronellal + + − 5 1.49 0.59 InhibitionofHIV-1protease.
OcimumtenuiflorumL. Carvacrol − − + 3 0.75 0.35 Inactivation ofviral protease incase of HIVinfection.
PaeonialactifloraPall. Paeoniflorin + − − 3 1.41 0.56 Inhibits viralentry in case ofInfluenza virusinfection.
PhyllanthusamarusSchumach.&Thonn.
Gallotannin − + + 5 1.49 0.59 Halts theprocess of viralreplication incase of Herpessimplex virusinfection.
RheumrhabarbarumL. Malicacid + − + 4 1.91 0.76 Inhibits viralentrybyceasingthe endosomalfusion incaseofinfluenzavirus.
SalviamiltiorrhizaBunge
Salvianolicacid − + + 5 1.49 0.59 InhibitionofHIV-1 integrase andprotease.
Taxillussutchuenensisvar.duclouxii(Lecomte)H.S.Kiu
Butenolide + − + 2 1.5 0.60 Inhibition ofHepatitis C viralNS3 serineprotease andceasing viral
entry.Tinosporacordifolia(Willd.)Hook.f.&Thomson
Tinosporaside + + + 5 2.49 1 Immunomodulatory activity;Anti-HIV activitywherein it acts
24
wherein it actsas viralribonucleaseinhibitor.
Withaniasomnifera(L.)Dunal
Withanolide + + + 3 1.41 0.56 Disruptsinteractionsbetween viral S-protein receptorbinding domainand Host ACE2receptor.
ZingiberofficinaleRoscoe
6-Gingerol − + + 5 1.49 0.59 InhibitsHepatitisCvirusprotease.
§Plants are selectedon thebasis of extensive literature surge, specifically focusingon their ethno-medicinalattributes,symptomaticreliefprovisionabilitiesanddirect/indirectantiviralactivity,ifany.†Symbolsof+and–denotethepresenceandabsenceofviralvirulencefactorinhibitorypropertiesinthe given plant, as deduced on the basis of keyword searchmatrix analysis using PubMed searchengine.*Fuzzy score μS= (S-minS)/ (maxS-minS), wherein shaded cells represent the ligands selected forfurtherstudywithafuzzyscore>0.5.
Table 4. Physicochemical properties of phytoligands in comparison with the standardchemotherapeuticagent.
Ligand/StandardPhysicochemicalProperties
Mol.Wt.(≤500D)
LogP(≤5)† H-BondDonor(≤5)
H-DonorAcceptor(≤10)
Lipinskiviolations(ifany)*
Allicin 162.02 0.237 0 1 0Amentoflavone 538.09 2.030 6 10 1
Berberine 336.12 2.473 0 4 0Beta-caryophyllene 204.19 6.044 0 0 111-keto-beta-boswellicacid
470.34 8.131 2 4 1
Butenolide 84.02 0.308 0 2 0Citronellal 154.14 3.591 0 1 0Curcumin 368.13 1.945 2 6 0Gallotannin 1700 9.537 25 46 4
Gamma-Glutamyl-S-allylcysteine
290.09 -2.68 4 7 0
6-Gingerol 294.18 2.437 2 4 0Malicacid 134.02 -1.474 3 5 0β-myrcene 136.13 4.170 0 0 0Paeoniflorin 480.16 -0.464 5 11 1Quercetin 302.04 1.834 5 7 0Salvianolicacid 494.12 2.898 7 10 1Tinosporaside 492.20 0.54 4 10 0Withanolide 470.27 3.263 2 6 0Hydroxychloroquine 335.88 4.00 4 2 0
†Logarithmofcompoundpartitioncoefficientbetweenn-octanolandwater.*Shadedcellindicatesphytoligandwithmorethan1Lipinskiviolationsandhenceiseliminatedatthis
stageitself.
Table5.ADMEToxvaluesofphytoligandsincomparisonwiththestandardchemotherapeuticagent.
25
Ligand/Standard
Absorption Distribution Metabolism Excretion
Caco-2permeability(value≥0.5)
Humanintestinalabsorption(value≥0.5)
PlasmaProteinbinding(value≥0.5)
Watersolubility(logS≥-4)
P-glyco-proteinactivator(value≥0.5)
CYP3A4inhibition(value≥0.5)
Acuteoraltoxicity(Kg/mol)(value≥1.0)
Allicin 0.58(+) 0.91(+) 0.50(+) -0.89(+) 0.98(+) 0.92(-) 1.935(-)Amentoflavone 0.87(+) 0.98(+) 1.11(+) -3.36(+) 0.44(-) 0.61(-) 1.822(-)
Berberine 0.94(+) 0.77(+) 0.83(+) -2.97(+) 0.68(+) 0.58(-) 1.545(-)
Beta-caryophyllene
0.86(+) 0.98(+) 0.83(+) -4.68(+) 0.89(+) 0.86(-) 2.366(-)
11-keto-beta-boswellicacid
0.54(+) 0.99(+) 1.05(+) -3.45(+) 0.63(+) 0.79(-) 2.834(-)
Butenolide 0.76(+) 0.96(+) 0.096(-) 0.23(+) 0.98(+) 0.98(-) 1.976(-)
Citronellal 0.92(+) 0.97(+) 0.70(+) -2.44(+) 0.98(+) 0.96(-) 2.307(-)
Curcumin 0.76(+) 0.97(+) 0.83(+) -3.36(+) 0.59(+) 0.53(-) 1.992(-)
Gamma-Glutamyl-S-allylcysteine
0.92(+) 0.63(+) 0.50(+) -1.68(+) 0.93(+) 0.74(-) 1.648(-)
6-Gingerol 0.59(+) 0.99(+) 0.85(+) -3.23(+) 0.89(+) 0.59(-) 2.290(-)
Malicacid 0.95(+) 0.77(+) 0.23(-) 0.27(+) 0.98(+) 0.90(-) 0.844(+)
β-myrcene 0.77(+) 0.96(+) 0.43(-) -3.44(+) 0.98(+) 0.66(-) 1.660(-)
Paeoniflorin 0.82(+) 0.41(-) 0.67(+) -2.97(+) 0.65(+) 0.85(-) 3.502(-)
Quercetin 0.64(+) 0.98(+) 1.17(+) -2.99(+) 0.91(+) 0.69(-) 2.559(-)
Salvianolicacid 0.93(+) 0.96(+) 1.03(+) -3.20(+) 0.65(+) 0.83(-) 2.069(-)
Tinosporaside 0.84(+) 0.83(+) 0.50(+) -3.65(+) 0.54(+) 0.75(-) 3.236(-)
Withanolide 0.62(+) 0.97(+) 1.18(+) -4.00(+) 0.51(+) 0.85(-) 3.660(-)
Hydroxy-chloroquine
0.66(+) 0.99(+) 0.86(+) -4.00(+) 0.84(+) 0.83(-) 2.684(-)
*Denoted ‘+’ or ‘-’ sign relates to the presence or absence of a predicted activity, respectively.Shadedcellsindicatethedescriptorsviolatingthestandardvalues,therebyexcludingtherespectivephytoligand(s)fromfurtherstudies.
Duetotechnicallimitations,table6isonlyavailableasadownloadinthesupplemental
filessection.
Figures
26
Figure1
MolecularPathogenesis&ClinicalmanifestationsofNovelCoronavirus(SARS-CoV-2).SARS-
CoV-2infectslungparenchymalcellsanditsentryisfacilitatedbybindingofviralenvelope
glycoprotein(S2)withhostACE2receptor.Uponenteringthehostbronchialcells,thevirus
elicitsanimmunologicalresponseasmediatedbyhostgenes,namely,TLRs-dependentIFN
inductionpathways[IRF-3/7(1)&NF-Kβ(2)]andmyeloid-differentiationprimaryresponse
88pathways[ATF-2(3)&AP-1(4)].Subsequently,inflammatorycytokines(IL-6,IL-12and
IL-8),interferongammaandreactiveoxygenspecies(ROS)areproduced.These
inflammatoryandoxidationmediatorsactivateapathogenesiscascade,leadingto
inflammation,alveolarmucusformationanddecreaseinpartialpressureofoxygenin
bronchioles.SimultaneouslySARS-CoV-2getsholdofhostmetabolicmachineryandinitiates
itsreplicationandtranslation,asmediatedbytheprimaryviralvirulencefactors(RNA
27
dependentRNApolymerase/endoribonuclease,andviralmainprotease).Asaconsequence,
viralloadinbloodincreases(viraemia),ultimatelyleadingtoseriousclinicalmanifestations
includingfever,alveolaredema,hypoxemia,dyspnoea,coughandmultipleorganfailure.
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Figure2
RamachandranplotofthestructuremodelsofSARS-CoV-2receptors-(A)Spike
glycoprotein,(B)NSP15endoribonuclease,and(C)Mainprotease3CLpro.Themost
favouredregionsarecolouredinredandmarkedasA,B,andL.Theadditionallyallowed
regionsarecolouredinyellowandmarkedasa,b,l,andp.Allnon-glycineandproline
residuesareshownasfilledblacksquares,whereasglycineresidues(non-end)areshown
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asfilledblacktriangles.Disallowedresidues,ifanyarerepresentedbywhitecolour.
Abbreviations:Asn:asparagine;Asp:aspartate;Gly:glycine;Pro:proline.
Figure3
Bindingposedepictionsofgamma-glutamyl-S-allylcysteineasprobabledrugcandidates
targetingSARS-CoV-2receptorsAmidegroupofgamma-glutamyl-S-allylcysteine(GGA)
formedahydrogenbondwiththeamideresidueofglutamineaminoacid(1071stposition)
oftheviralspikeglycoprotein.Glutamicacidresidues(45thposition)ofviral
endoribonucleaseshowedirreversibleinteractionwithGGA.Additionally,thehydroxylgroup
ofGGAformedahydrogenbondwithcarbonylgroupofprolineaminoacid(108thposition)
ofviralmainprotease3CLpro.
SupplementaryFilesThisisalistofsupplementaryfilesassociatedwiththispreprint.Clicktodownload.
GraphicalAbstract.jpgTable2.docxTable6.docx
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