Scyllatoxin(Leiurotoxin-1)isaneurotoxinthatwasoriginallyisolatedfromLeiurusquinquestriatushebraeus.ScyllatoxinbindsandblocksSKchannels(smallconductanceCa2+-activatedK+channels)inthebrainandspinalcordandinhibitsthem.
Description:
AAsequence:Ala-Phe-Cys3-Asn-Leu-Arg-Met-Cys8-Gln-Leu-Ser-Cys12-Arg-Ser-Leu-Gly-Leu-Leu-Gly-Lys-Cys21-Ile-Gly-Asp-Lys-Cys26-Glu-Cys28-Val-Lys-His-NH2
Disulfidebondsbetween:Cys3-Cys21,Cys8-Cys26andCys12-Cys28
Length(aa):31
Formula:C142H237N45O39S7
MolecularWeight:3424.1Da
Appearance:Whitelyophilizedsolid
Solubility:waterandsalinebuffer
CASnumber:[116235-63-3]Source:Synthetic
Purityrate:>95%
Reference:
EffectofcharyBDotoxinandleiurotoxinIonpotassiumcurrentsinbullfrogsympatheticganglionandhippocampalneurons
TheeffectsofcharybdotoxinandleiurotoxinIwereexaminedonseveralclassesofK+currentsinbullfrogsympatheticganglionandhippocampalCA1pyramidalneurons.Highlypurifiedpreparationsofcharybdotoxinselectivelyblockedalargevoltage-andCa(2+)-dependentK+current(IC)responsIBLeforactionpotentialrepolarization(IC50=6nM)whileleiurotoxinIselectivelyblockedasmallCa(2+)-dependentK+conductance(IAHP)responsiblefortheslowafterhyperpolarizationfollowinganactionpotential(IC50=7.5nM)inbullfrogsympatheticganglionneurons.NeitherofthetoxinshadsignificanteffectsonotherK+currents(M-current[IM],A-current[IA]andthedelayedrectifier[IK])presentinthesecells.LeiurotoxinIataconcentrationof20nMhadnodetectableeffectoncurrentsinhippocampalCA1pyramidalneurons.ThislackofeffectonIAHPincentralneuronssuggeststhatthechannelsunderlyingslowAHPsinthoseneuronsarepharmacologicallydistinctfromanalogouschannelsinperipheralneurons.
GohJW.,etal.(1992)EffectofcharybdotoxinandleiurotoxinIonpotassiumcurrentsinbullfrogsympatheticganglionandhippocampalneurons.BrainRes.PMID: 1280181
Anapamin-andscyllatoxin-insensitiveisoformofthehumanSK3channel
WehaveisolatedanhSK3isoformfromahumanembryonicCDNAlibrarythatwehavenamedhSK3_ex4.Thisisoformcontainsa15aminoacidinsertionwithintheS5toP-loopsegment.TranscriptsencodinghSK3_ex4arecoexpressedatlowerlevelswithhSK3inneuronalaswellasinnon-neuronaltissues.ToinvestigatethepharmacokineticpropertiesofhSK3_ex4,weexpressedtheisoformshSK3andhSK3_ex4intsAcells.BothisoformsweresimilarlyactivatedbycytosolicCa2+(hSK3,EC50=0.91+/-0.4microM;hSK3_ex4,EC50=0.78+/-0.2microM)andby1-ethyl-2-benzimidazolinone(hSK3,EC50=0.17mM;hSK3_ex4,0.19mM).Theywerebothblockedbytetraethylammonium(hSK3,Kd=2.2mM;hSK3_ex4,2.6mM)andshowedsimilarpermeABIlitiesrelativetoK+forCs+(hSK3,0.17+/-0.04,n=3;hSK3_ex4,0.17+/-0.05,n=3)andRb+(hSK3,0.79+/-0.04,n=3;hSK3_ex4,0.8+/-0.07,n=3).Ba2+blockedbothisoforms,andinbothcases,theblockwasstrongestathyperpolarizingmembranepotentials.However,thevoltage-dependenceofhSK3wasstrongerthanthatofhSK3_ex4.ThemostobviousdistinguishingfeatureofthisnewisoformwasthatwhereashSK3wasblockedbyapamin(Kd=0.8nM),scyllatoxin(Kd=2.1nM),andd-tubocurarine(Kd=33.4microM),hSK3_ex4wasnotaffectedbyapaminupto100nM,scyllatoxinupto500nM,andd-tubocurarineupto500microM.Sofar,isoformhSK3_ex4formstheonlysmall-conductancecalcium-activatedpotassium(SK)channels,whichareinsensitivetotheclassicSKblockers.
WittekindtOH.,etal.(2004)Anapamin-andscyllatoxin-insensitiveisoformofthehumanSK3channel.MolPharmacol.PMID: 14978258
LeiurotoxinI,ascorpiontoxinspecificforCa(2+)-activatedK+channels.Structure-activityanalysisusingsyntheticanalogs
Recently,wereportedastructure-activityrelationshipstudyonP05,anovelleiurotoxinI-likescorpiontoxinwhichisselectivefortheapamin-sensitiveCa(2+)-activatedK+channel[Sabatieretal.(1993)Biochemistry32,2763-2770].Arg6,Arg7andC-terminalHis31appearedtobekeyresiduesforP05BIOLOGicalactivity.OwingtothehighsequenceidentitybetweenP05andleiurotoxinI(87%),severalanalogsofleiurotoxinI(Lei-NH2)withpointmutationsatthesepositionsweredesignedandchemicallysynthesizedusinganoptimizedsolid-phasetechnique.Thesynthesizedpeptideswere[L6]Lei-NH2,[R7]Lei-NH2,Lei-OHand[R7]Lei-OH,aswellasfragment[R7,Abu8]N4-S11-NH2.Achimericanalog([M22,K24,R27]Lei-NH2),whichpossessespartoftheiberiotoxinC-terminus,wasalsoconstructed.Circulardichroismanalysesoftheseanalogs,inagreementwiththeirstructuralmodelsobtainedbymoleculardynamics,showedthatthepointmutationsdidnotsignificantlyaffecttheoverallsecondarystructures,ascomparedtonaturalLei-NH2.Allthepeptidesandnaturaltoxinswerecomparedinvitrofortheircapacitytoinhibitbindingof[125I]-apamintoratbrainsynaptosomes,andinvivofortheirspecificneurotoxicityinmice.TheArg6residuewasessentialforhighbiologicalactivityofleiurotoxinI.Further,substitutionofMet7inthenaturaltoxinbyArg7,orC-terminalamidationofHis31,greatlyincreasedaffinityfortheapaminreceptorbutdidnotsignificantlyaffecttoxinneurotoxicity.Remarkably,thechimericanalog[M22,K24,R27]Lei-NH2wasfoundtoretainleiurotoxinI-likeactivity,thusindicatingthatthenegativelychargedresiduesAsp24andGlu27(andIle22)arenotdirectlyinvolvedinthehightoxinbioactivity.However,thechimericmoleculehadnoiberiotoxin-likeeffectonratmuscularmaxi-K+channelsincorporatedinlipidbilayers.
SabatierJM.,etal.(1994)LeiurotoxinI,ascorpiontoxinspecificforCa(2+)-activatedK+channels.Structure-activityanalysisusingsyntheticanalogs.IntJPeptProteinRes. PMID: 8070973
Scyllatoxin,ablockerofCa(2+)-activatedK+channels:structure-functionrelationshipsandbrainlocalizationofthebindingsites
Chemicalmodificationsofscyllatoxin(leiurustoxinI)haveshownthattwoargininesinthesequence,Arg6andArg13,areessentialbothforbindingtotheCa(2+)-activatedK+channelproteinandforthefunctionaleffectofthetoxin.His31isimportantbothforthebindingactivityofthetoxinandfortheinductionofcontractionsontaeniacoli.However,althoughitsiodinationdrasticallydecreasesthetoxinactivity,itdoesnotabolishit.ChemicalmodificationoflysineresiduesorofGlu27doesnotsignificantlyaltertoxinbinding,butitdrasticallydecreasespotencywithrespecttocontractionoftaeniacoli.Thesameobservationhasbeenmadeafterchemicalmodificationofthelysineresidues.ThebraindistributionofscyllatoxinbindingsiteshasbeenanalyzedbyquantitativeautorADIographicanalysis.Itindicatesthatapamin(abeevenomtoxin)bindingsitesarecolocalizedwithscyllatoxinbindingsites.Theresultsareconsonantwiththepresenceofapamin/scyllatoxinbindingsitesassociatedwithCa(2+)-activatedK+channels.High-affinitybindingsitesforapamincanbeassociatedwithvery-high-affinity(lessthan70pM),high-affinity(approximately100-500pM),ormoderate-affinity(greaterthan800pM)bindingsitesforscyllatoxin.
AugusteP.,etal.(1992)Scyllatoxin,ablockerofCa(2+)-activatedK+channels:structure-functionrelationshipsandbrainlocalizationofthebindingsites.Biochemistry. PMID: 1731919
Purificationandcharacterizationofaunique,potentinhibitorofapaminbindingfromLeiurusquinquestriatushebraeusvenom
Aninhibitorofapaminbindinghasbeenpurifiedtohomogeneityinthreechromatographicstepsfromthevenomofthescorpion,Leiurusquinquestriatushebraeus.Theinhibitor,whichwehavenamedleiurotoxinI,representslessthan0.02%ofthevenomprotein.Itisa3.4-kDapeptidewithlittlestructuralhomologytoapaminalthoughithassomehomologytootherscorpiontoxinssuchascharybdotoxin,noxiustoxin,andneurotoxinP2.LeiurotoxinIcompletelyinhibits125I-apaminbindingtoratbrainsynaptosomalmembranes(Ki=75pM).Thus,itis10-20-foldlesspotentthanapamin.LeiurotoxinIisnotastrictlycompetitiveinhibitorofthisbindingreaction.Likeapamin,leiurotoxinIblockstheepinephrine-inducedrelaxationofguineapigteniaecoli(ED50=6.5nM),whilehavingnoeffectontherateorforceofcontractioninguineapigatriaorrabbitportalveinpreparations.Thus,leiurotoxinIofscorpionvenomandapaminofhoneybeevenomdemonstratesimilaractivitiesinavarietyoftissues,yetarestructurallyunrelatedpeptides.Thesetwopeptidesshouldbeusefulinelucidatingtheroleofthesmallconductance,Ca2+-activatedK+channelsindifferenttissues.
ChicchiGG.,etal.(1988)Purificationandcharacterizationofaunique,potentinhibitorofapaminbindingfromLeiurusquinquestriatushebraeusvenom. PMID: 2839478
Iberiotoxin-sensitiveand-insensitiveBKcurrentsinPurkinjeneuronsomata
Purkinjecellshavespecializedintrinsicionicconductancesthatgeneratehigh-frequencyactionpotentials.DisruptionsoftheirCaorCa-activatedK(KCa)currentscorrelatewithalteredfiringpatternsinvitroandimpairedmotorbehaviorinvivo.ToexaminethepropertiesofsomaticKCacurrents,werecordedvoltage-clampedKCacurrentsinPurkinjecellbodiesisolatedfrompostnatalday17-21mousecerebellum.CurrentswereevokedbyendogenousCainfluxwithapproximatelyphysiologicalCabuffering.Purkinjesomataexpressedvoltage-activated,Cd-sensitiveKCacurrentswithiberiotoxin(IBTX)-sensitive(>100nS)andIBTX-insensitive(>75nS)components.IBTX-sensitivecurrentsactivatedandpartiallyinactivatedwithinmilliseconds.Rapid,incompletemacroscopicinactivationwasalsoevidentduring50-or100-Hztrainsof1-msdepolarizations.Incontrast,IBTX-insensitivecurrentsactivatedmoreslowlyanddidnotinactivate.Thesecurrentswereinsensitivetothesmall-andintermediate-conductanceKCachannelblockersapamin,scyllatoxin,UCL1684,bicucullinemethiodide,andTRAM-34,butwerelargelyblockedby1mMtetraethylammonium.Theunderlyingchannelshadsingle-channelconductancesof∼150pS,suggestingthatthecurrentsarecarriedbyIBTX-resistant(β4-containing)large-conductanceKCa(BK)channels.IBTX-insensitivecurrentswereneverthelessincreasedbysmall-conductanceKCachannelagoNISTsEBIO,chlorzoxazone,andCyPPA.Duringtrainsofbriefdepolarizations,IBTX-insensitivecurrentsflowedduringinterstepintervals,andtheaccumulationofinterstepoutwardcurrentwasenhancedbyEBIO.Incurrentclamp,EBIOslowedspiking,especiallyduringdepolarizingcurrentinjections.ThetwocomponentsofBKcurrentinPurkinjesomatalikelycontributedifferentlytospikerepolarizationandfiringrate.Moreover,augmentationofBKcurrentmaypartiallyunderlietheactionofEBIOandchlorzoxazonetoalleviatedisruptedPurkinjecellfiringassociatedwithgeneticataxias.
BentonMD.,etal.(2013)Iberiotoxin-sensitiveand-insensitiveBKcurrentsinPurkinjeneuronsomata.JNeurophysiol. PMID: 23446695
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