FluorescentNav1.7blocker
Cy5-ProTx-IIisafluorescentlylabeledProTx-II,afamousNav1.7blocker.ThewildtypeProTx-IIblocksNav1.7 withanIC50 valueofaround300pM,Nav1.2, Nav1.5and Nav1.6 withIC50 valuesof41nM,79nMand26nMrespectively.TheCy5-ProTx-IIversiondevelopedbySmartoxhaspotentNav1.7blockingactivity.ItwasshowntofullyblockNav1.7at100nMconcentration.
A,Recordingtracesoftransiently-expressedhumanNav1.7currentinthepresenceofCy5-ProTx-II(100nM).Thecurrentwaselicitedbya50ms-depolarizingpulseto-10mVfromaholdingpotentialof-90mV.Inter-sweepperiodwas10s.Currentamplitudeswereplottedagainsttime.Notethattoxin-inducedinhibitionisresistanttowashout,howeveritcanbepartiallyrelievedbydepolarizingthecellmembraneatthetimepointindicatedbytheredbar.B,C,RecordingtracesofhNav1.7currentinthepresenceofCy5-ProTx-II(100nM)fromtwodistinctcells.Membranedepolarizationfailstorelievetheinhibitioninthepresenceof100nMCy5-ProTx-II;howevertheinhibitioncanbepartiallyrelievedbydepolarizationafterwashout.D,FamiliesofhNav1.7currenttracesincontrolandinthepresenceof100nMCy5-ProTx-II.Currentswereevokedbydepolarizingpulsesfrom-60mVto40mV,whilethecellwasholdat-90mV.E,Amplitude-voltagerelationshipsobtainedfromD.
Description:
AAsequence: Tyr-Cys2-Gln-Lys-Trp-Met-Trp-Thr-Cys9-Asp-Ser-Glu-Arg-Lys-Cys15-Cys16-Glu-Gly-Met-Val-Cys21-Arg-Leu-Trp-Cys25-Lys-Lys-Lys-Leu-Trp-OH
Disulfidebonds: Cys2-Cys16,Cys9-Cys21,Cys15-Cys25
Length(aa): 30
ProTx-IIformula: C168H250N46O41S8
MolecularWeight:closeto4450g/mol
Appearance:darklyophilizedsolid
Source: Synthetic
Purityrate: >95%
Cy5:λex646nm,λem662nm
Stoichiometry:1:1
Reference:
Spider-venompeptidesthattargetvoltage-gatedsodiumchannels:pharmacologicaltoolsandpotentialtherapeuticleads
Voltage-gatedsodium(Na(V))channelsplayacentralroleinthepropagationofactionpotentialsinexcitablecellsinbothhumansandinsects.ManyvenomousanimalshavethereforeevolvedtoxinsthatmodulatetheactivityofNa(V)channelsinordertosuBDuetheirpreyanddeterpredators.SpidervenomsinparticulararerichinNa(V)channelmodulators,withone-thirdofallknownionchanneltoxinsfromspidervenomsactingonNa(V)channels.Herewereviewthelandscapeofspider-venompeptidesthathavesofarbeendescribedtotargetvertebrateorinvertebrateNa(V)channels.Thesepeptidesfallinto12distinctfamiliesbasedontheirprimarystructureandcysteinescaffold.Someofthesepeptideshavebecomeusefulpharmacologicaltools,whileothershavepotentialastherapeuticleadsbecausetheytargetspecificNa(V)channelsubtypesthatareconsideredtobeimportantanalgesictargets.Spidervenomsareconservativelypredictedtocontainmorethan10millionbioactivepeptidesandsofaronly0.01%ofthisdiversitybeencharacterised.Thus,itislikelythatfutureresearchwillrevealadditionalstructuralclassesofspider-venompeptidesthattargetNa(V)channels.
KlintJK., etal. (2012)Spider-venompeptidesthattargetvoltage-gatedsodiumchannels:pharmacologicaltoolsandpotentialtherapeuticleads. Toxicon. PMID:22543187
EvidenceformultipleeffectsofProTxIIonactivationgatinginNa(V)1.5
ThepeptidetoxinProTxII,recentlyisolatedfromthevenomofthetarantulaspiderThrixopelmapruriens,modifiesgatinginvoltage-gatedNa+andCa2+channels.ProTxIIisdistinctfromotherknownNa+channelgatingmodifiertoxinsinthatitaffectsactivation,butnotinactivation.Itshiftsactivationgatingpositivelyanddecreasescurrentmagnitudesuchthatthedose-dependenceoftoxinactionmeasuredatasinglepotentialreflectsbotheffects.Totesttheextenttowhichtheseeffectswereindependent,wetrackedseveraldifferentmeasuresofcurrentamplitude,voltage-dependentactivation,andcurrentkineticsinNa(V)1.5inarangeoftoxinconcentrations.ChangesinvoltagedependenceandadecreaseinG(max)appearedatrelativelylowconcentrations(40-100nM)whileapositiveshiftinthevoltagerangeofactivationwasapparentathighertoxinconcentrations(>or=500nM).BecauseProTxIIcarriesanet+4chargewetestedwhetherelectrostaticinteractionscontributedtotoxinaction.WeexaminedtheeffectsofProTxIIinthepresenceofhighextracellularBa2+,knowntoscreenand/orbindtosurfacecharge.Some,butnotallaspectsofProTxIImodificationweresensitivetothepresenceofBa2+indicatingthecontributionofanelectrostatic,surfacecharge-likemechanismandsupportingtheideaofamulti-facetedtoxin-channelinteraction.
EdgertonG.B., etal. (2008)EvidenceformultipleeffectsofProTxIIonactivationgatinginNa(V)1.5, Toxicon. PMID:18657562
ProTx-II,aselectiveinhibitorofNav1.7sodiumchannels,blocksactionpotentialpropagationinnociceptors
Voltage-gatedsodium(Na(V)1)channelsplayacriticalroleinmodulatingtheexcitABIlityofsensoryneurons,andhumangeneticevidencepointstoNa(V)1.7asanessentialcontributortopainsignaling.Humanloss-of-functionmutationsinSCN9A,thegeneencodingNa(V)1.7,causechannelopathy-associatedindifferencetopain(CIP),whereasgain-of-functionmutationsareassociatedwithtwoinheritedpainfulneuropathies.AlthoughthehumangeneticdatamakeNa(V)1.7anattractivetargetforthedevelopmentofanalgesics,pharmacologicalproof-of-conceptinexperimentalpainmodelsrequiresNa(V)1.7-selectivechannelblockers.Here,weshowthatthetarantulavenompeptideProTx-IIselectivelyinteractswithNa(V)1.7channels,inhibitingNa(V)1.7withanIC(50)valueof0.3nM,comparedwithIC(50)valuesof30to150nMforotherheterologouslyexpressedNa(V)1subtypes.ThissubtypeselectivitywasabolishedbyapointmutationinDIIS3.ItisinterestingthatapplicationofProTx-IItodesheathedcutaneousnervescompletelyblockedtheC-fibercompoundactionpotentialatconcentrationsthathadlittleeffectonAbeta-fiberconduction.ProTx-IIapplicationhadlittleeffectonactionpotentialpropagationoftheintactnerve,whichmayexplainwhyProTx-IIwasnotefficaciousinrodentmodelsofacuteandinflammatorypain.Mono-iodo-ProTx-II((125)I-ProTx-II)bindswithhighaffinity(K(d)=0.3nM)torecombinanthNa(V)1.7channels.Bindingof(125)I-ProTx-IIisinsensitivetothepresenceofotherwellcharacterizedNa(V)1channelmodulators,suggestingthatProTx-IIbindstoanovelsite,whichmaybemoreconducivetoconferringsubtypeselectivitythanthesiteoccupiedbytrADItionallocalanestheticsandanticonvulsants.Thus,the(125)I-ProTx-IIbindingassay,describedhere,offersanewtoolinthesearchfornovelNa(V)1.7-selectiveblockers.
WilliamA., etal.(2007)ProTx-II,aselectiveinhibitorofNav1.7sodiumchannels,blocksactionpotentialpropagationinnociceptors. Mol.Pharm. PMID:18728100
ProTx-IandProTx-II:gatingmodifiersofvoltage-gatedsodiumchannels
ThetarantulavenompeptidesProTx-IandProTx-IIinhibitvoltage-gatedsodiumchannelsbyshiftingtheirvoltagedependenceofactivationtoamorepositivepotential,thusactingbyamechanismsimilartothatofpotassiumchannelgatingmodifierssuchashanatoxinandVSTX1.ProTx-IandProTx-IIinhibitallsodiumchannel(Nav1)subtypestestedwithsimilarpotencyandrepresentthefirstpotentpeptidylinhibitorsofTTX-resistantsodiumchannels.Likegatingmodifiersofpotassiumchannels,ProTx-IandProTx-IIconformtotheinhibitorycystineknotmotif,andProTx-IIwasdemonstratedtobindtosodiumchannelsintheclosedstate.Bothtoxinshavebeensynthesizedchemically,andProTx-II,producedbyrecombinantmeans,hasbeenusedtomaptheinteractionsurfaceofthepeptidewiththeNav1.5channel.Incomparison,beta-scorpiontoxinsactivatesodiumchannelsbyshiftingthevoltagedependenceofactivationtomorenegativepotentials,andtogetherthesepeptidesrepresentvaluabletoolsforexploringthegatingmechanismofsodiumchannels.
PriestB.T., etal.(2007)ProTx-IandProTx-II:gatingmodifiersofvoltage-gatedsodiumchannels, Toxicon. PMID:17087985
Differentialphospholipidbindingbysite3andsite4toxins.Implicationsforstructuralvariabilitybetweenvoltage-sensitivesodiumchanneldomains
Ithasbeenshownrecentlythatpolypeptidetoxinsthatmodulatethegatingpropertiesofvoltage-sensitivecationchannelsareabletobindtophospholipidmembranes,leadingtothesuggestionthatthesetoxinsareabletoaccessachannel-bindingsitethatremainsmembrane-restricted(Lee,S.-Y.,andMacKinnon,R.(2004)Nature430,232-235).WethereforeexaminedtheabilityofanthopleurinB(ApB),aseaanemonetoxinthatselectivelymodifiesinactivationkineticsofNa(V)1.xchannels,andProTx-II,aspidertoxinthatmodifiesactivationkineticsofthesamechannels,tobindtoliposomes.WhereasProTx-IIcanbequantitativelydepletedfromsolutionuponincubationwithphosphatidylcholine/phosphatidylserineliposomes,ApBdisplaysnodiscernIBLephospholipidbindingactivity.Wethereforeexaminedtheactivitiesofstructurallyunrelatedsite3andsite4toxinsderivedfromLeiurusandCentruroidesvenoms,respectively,inthesameassay.LikeApB,thesite3toxinLqqVshowsnolipidbindingactivity,whereasthesite4toxinCentruroidestoxinII,likeProTx-II,iscompletelybound.WeconcludethattoxinsthatmodifyinactivationkineticsviabindingtoNa(V)1.xsite3lacktheabilitytobindphospholipids,whereassite4toxins,whichmodifyactivation,havethisactivity.ThisinherentdifferencesuggeststhattheconformationofdomainIImorecloselyresemblesthatoftheK(V)APchannelthandoestheconformationofdomainIV.
SmithJ.J., etal. (2005)Differentialphospholipidbindingbysite3andsite4toxins.Implicationsforstructuralvariabilitybetweenvoltage-sensitivesodiumchanneldomains, JBiolChem. PMID:15632158
Twotarantulapeptidesinhibitactivationofmultiplesodiumchannels
Twopeptides,ProTx-IandProTx-II,fromthevenomofthetarantulaThrixopelmapruriens,havebeenisolatedandcharacterized.Thesepeptideswerepurifiedonthebasisoftheirabilitytoreversiblyinhibitthetetrodotoxin-resistantNachannel,Na(V)1.8,andareshowntobelongtotheinhibitorycystineknot(ICK)familyofpeptidetoxinsinteractingwithvoltage-gatedionchannels.Thefamilyhasseveralhallmarks:cystinebridgeconnectivity,mechanismofchannelinhibition,andpromiscuityacrosschannelswithinandacrosschannelfamilies.ThecystinebridgeconnectivityofProTx-IIisverysimilartothatofothermembersofthisfamily,i.e.,C(2)toC(16),C(9)toC(21),andC(15)toC(25).Thesepeptidesarethefirsthigh-affinityligandsfortetrodotoxin-resistantperipheralnerveNa(V)channels,butalsoinhibitotherNa(V)channels(IC(50)’s<100nM).ProTx-IandProTx-IIshiftthevoltagedependenceofactivationofNa(V)1.5tomorepositivevoltages,similartoothergating-modifierICKfamilymembers.ProTx-IalsoshiftsthevoltagedependenceofactivationofCa(V)3.1(alpha(1G),T-type,IC(50)=50nM)withoutaffectingthevoltagedependenceofinactivation.Toenablefurtherstructuralandfunctionalstudies,syntheticProTx-IIwasmade;itadoptsthesamestructureandhasthesamefunctionalpropertiesasthenativepeptide.SyntheticProTx-Iwasalsomadeandexhibitsthesamepotencyasthenativepeptide.SyntheticProTx-I,butnotProTx-II,alsoinhibitsK(V)2.1channelswith10-foldlesspotencythanitspotencyonNa(V)channels.ThesepeptidesrepresentnoveltoolsforexploringthegatingmechanismsofseveralNa(V)andCa(V)channels.
MiddletonR.E., etal. (2002)Twotarantulapeptidesinhibitactivationofmultiplesodiumchannels, Biochemistry. PMID:12475222
Smartox Biotechnolgy的多肽毒素产品如下：
1. 作用于钠离子通道(Sodium channel)的毒素
Toxin name | Catalog # | Target |
Phrixotoxin-3 | 13PHX003 | Selective blocker of Nav1.2 |
µ-conotoxin GIIIB | CON020 | Selective blocker of Nav1.4 |
µ-conotoxin CnIIIC | CON021 | Selective blocker of Nav1.4 |
μ-conotoxin PIIIA | 08CON006 | Selective blocker of Nav1.4 |
Jingzhaotoxin-III | 12JZH003 | Selective blocker of Nav1.5 |
ProTx-II | 07PTX002 | Selective blocker of Nav1.7 |
ProTx-II Biotin | 12PTB002 | Selective blocker of Nav1.7 |
ProTx-I | 12PTX001 | Blocker of Nav1.8, Nav1.2, Nav1.5, Nav1.7 |
Huwentoxin-I | 07HWT001 | Blocker of TTX-S |
Huwentoxin-IV | 08HWT002 | Blocker of TTX-S |
Hainantoxin-III | 13HTX003 | Blocker of TTX-S |
Hainantoxin-IV | 12HTX001 | Blocker of TTX-S |
GsAF-I | 12GSF001 | Blocker of TTX-S |
GsAF-II | 12GSF002 | Blocker of TTX-S |
2. 作用于钾离子通道(Potassium channel)的毒素
Toxin name | Catalog # | Target |
KCa channels | ||
Apamin 蜜蜂神经毒素 | 08APA001 | SK1, SK2, SK3 |
Charybdotoxin 蝎毒素 | 11CHA001 | KCa1.1, KCa3.1 - Kv1.2, Kv1.3, Kv1.6 |
Iberiotoxin | 12IBX001 | KCa1.1 |
Leiurotoxin 1 (Scyllatoxin) | 10LEI001 | SK1, SK2, SK3 |
Tamapin | 10TAM001 | SK1, SK2, SK3 |
Kaliotoxin-1 | 08KTX002 | BK, Kv1.1, Kv1.2, Kv1.3 |
Kv channels | ||
ShK | 08SHK001 | Kv1.3, Kv1.1, Kv1.4, Kv1.6 |
TMR-ShK | SAT001 | Kv1.3, Kv1.1 |
Margatoxin | 08MAG001 | Kv1.3 |
(Dap22)-ShK | 13SHD001 | Kv1.3 |
ADWX-1 | 13ADW001 | Kv1.3 |
HsTx1 | 08NEU001 | Kv1.3, Kv1.2 |
Agitoxin-2 | 13AGI002 | Kv1.3, Kv1.1 |
Maurotoxin | 08MAR001 | Kv1.2, KCa3.1 |
Guangxitoxin 1E | 11GUA002 | Kv2.1, Kv2.2 |
Stromatoxin 1 NEW | SCT01 | Kv2.1, Kv2.2 |
Kaliotoxin-1 | 08KTX002 | BK, Kv1.1, Kv1.2, Kv1.3 |
Charybdotoxin | 11CHA001 | KCa1.1, KCa3.1 - Kv1.2, Kv1.3, Kv1.6 |
Phrixotoxin-2 | PHX002 | Kv4.2, Kv4.3 |
AmmTx3 NEW | AMX001 | A-type potassium channels |
Inwardly rectifying potassium channels | ||
TertiapinQ | 08TER001 | Kir1.1, Kir3.1/3.4, Kir3.1/3.2-KCa1.1 |
hERG/Kv11.1 | ||
BeKm-1 | 13BEK001 | ERG1 |
3. 作用于钙离子通道(Calcium channel)的毒素
Toxin name | Catalog # | Target |
High voltage-gated Ca2+ channels | ||
ω-agatoxin IVA | 11AGA001 | P/Qtype |
ω-Conotoxin MVIIC | 08CON002 | P/Qtype, N-type |
ω-Conotoxin MVIIA | 08CON001 | N-type |
ω-Conotoxin GVIA | 08CON003 | N-type |
ω-Conotoxin SO3 | 08CON013 | N-type |
Huwentoxin I | 07HWT001 | N-type |
ProTx-II | 07PTX002 | T-type, L-type |
Intermediate voltage-gated Ca2+ channels | ||
SNX482 | 08SNX002 | R-type |
Low voltage-gated Ca2+ channels | ||
ProTx-I | 12PTX001 | T-type |
ProTx-II | 07PTX002 | T-type, L-type |
Ryanodine receptors | ||
Maurocalcine | 07PAU001 | Ryr1 |
4. 作用于氯离子通道(Chloride channel)的毒素
Toxin name | Catalog # | Target |
Chlorotoxin | 08CHL001 | Blocker of small conductance Cl- channels |
GaTx1 | 13GTX001 | Selective blocker of CFTR channel |
GaTx2 | 10GTX002 | Selective blocker of ClC-2 channel |
5. 作用于乙酰胆碱受体(Acetylcholine receptor)的毒素
Toxin name | Catalog # | Target |
α-conotoxin PeIA | 13CON017 | α9α10, α3β2 subunits |
α-Conotoxin PrXA | 13CON016 | α1/β1/ε/δ, α1/β1/γ/δ subunits |
Waglerin-1 | 12WAG001 | MusclenAChR |
α-conotoxin MI | 08CON012 | α1/δsubunits |
α-conotoxin GI | 08CON005 | α/δsite |
α-conotoxin IMI | 08CON011 | α7 homomeric nAChR |
α-conotoxin GID | CON019 | Blocker of α3β2, α7 and α4β2 nAChRs |
6. 含N-甲基-D-天冬氨酸NR2B
(NMDA, NR2B containing N-methyl-D-aspartate)
Conantokin-G
选择性、特异性抑制含NR2B的NMDAR。Conantokin-G能剂量依赖性抑制Ca2+内流，抑制NMDA诱导的兴奋性中毒效应。研究表明，在小鼠皮层神经元，Conantokin-G阻滞NMDA引发的电流信号的IC50值为480 nM。
7. 作用于酸敏感离子通道(ASIC channel, Acid-Sensing Ion Channel)的毒素
Toxin name | Catalog # | Target |
APETx2 | 07APE002 | Selective blocker of ASIC3 |
Psalmotoxin1/PcTx1 | 13PCT001 | Selective blocker of ASIC1a |
Ugr9-1 | 13UGR001 | Blocker of ASIC3 |
8. 作用于瞬时受体电位(TRP channel, transient receptor potential)的毒素
Toxin name | Catalog # | Target |
GsMTx4 | 08GSM001 | TRPC, TRPA |
Vanillotoxin3 | 10VAN003 | Activator of TRPV1 |
ProTx-I | 12PTX001 | Antagonist of TRPA1 |
9. 作用于嘌呤能通道(Purinergic channel)的毒素
Purotoxin-1
选择性抑制P2X3受体。100 nM Purotoxin-1 (PT-1)选择性抑制P2X3受体通道，在大鼠DRG神经元上，使用膜片钳实验表明：PT-1对电压门控通道和TRPV1均无抑制效应。10 µM ATP和100 µM α,β Methylene-ATP浓度下Purotoxin-1对P2X3受体有选择性作用，在该ATP浓度下Purotoxin-1对P2X2和杂化二聚体P2X2/3并无激动作用。Purotoxin-1对疼痛的潜在治疗作用。
10. 作用于其它膜受体通道(Others)的毒素
Smartox Biotechnology还提供其他类型的膜受体抑制剂：
Toxin name | Catalog # | Target |
Morphiceptin | 011CAS001 | Agonist of µ-opoid receptors |
Lys-conopressin G | 11CON14 | Vasopressin-like peptide |
GsMTx4 | 08GSM001 | Mechano sensitive ion channels |
Obtustatin | 10OBT001 | Blocks the binding of α1β1 integrin to collagen IV |
Rho-Conotoxin TIA | CON022 | Blocks α1-adrenergic receptor |
公司简介
Smartox Biotechnology是全球唯一一家专门生产动物毒液多肽毒素，用于细胞离子通道功能研究的生物医药公司。多肽毒素在生物制药领域具有重要的使用价值。
Smartox Biotechnology于2009年由来自Grenoble神经科学研究所(Grenoble Institute of Neuroscience)的Michel De Waard博士创立。Smartox Biotechnology专门研究动物毒液，制作合成多种毒液中的多肽成分（常称为毒素）。De Waard博士研究离子通道与毒素多肽的关系，尤其是鉴定、开发毒素多肽作为治疗性分子或细胞穿透肽(cell penetrating peptides, CPP)。其研究团队在毒液分离，药理性活性肽鉴定、富半胱氨酸肽定性、制作和优化等方面具有独特、丰富的经验。2010年，Smartox Biotechnolgy被法国研究部(Ministry of Research)授予“新兴企业OSEO奖(OSEO prize for emerging businesses)”。
总之，Smartox Biotechnolgy提供一系列高质量、具开创价值的多肽毒素。这些化合物在离子通道 研究中具有高的亲和性和选择性，是相应领域科学研究理想的生物毒素提供商和贴心的合作伙伴。
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