| Formulation | 25mMNaCitrate,100mMNaCl,100mMGlycine,pH6.8 |
| Storage | -80°C |
| Purity | >95%bySDS-PAGE |
| ActivityDetermination | N/A |
| ShelfLife(properlystored) | 12months |
Theformationofa1.8MDavonWillebrandfactor(vWF)multimerwithboundfactorVIIIisillustrated.EachvWFmonomer(Mr=260,000)containsafactorVIIIbindingsiteneartheNH2-terminalendofthemolecule.Themonomersarejoinedend-to-end(NH2toNH2andCOOHtoCOOH)bydisulfidebondstoformlargemultimers.ThematuremultimerscanbindonefactorVIIImoleculepermonomericsubunit.
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanvonWillebrandFactor,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
Overview:
VonWillebrandfactor(vWF)isamultimericplasmaglycoproteinthatisrequiredfornormalhemostaticplateletplugformation(1-8).Thematureplasmaproteiniscomposedofapparentlyidenticalsubunits(Mr=260,000)whichareheldtogetherbydisulfidebonds.ThecirculatingvWFmoleculerangesinsizefromdimers(Mr=520,000)toextremelylargemultimers(Mr>10,000,000).Duringnormalhemostasis,thelargermultimersofvWFareresponsIBLeforfacilitatingplateletplugformationbyformingabridgebetweenplateletglycoproteinIBandexposedcollageninthesubendothelium(9-14).EitheralackofvWFproteinorthepresenceofabnormalitieswhichresultindecreasedpolymerizationmaycausealossofBIOLOGicalactivitywhichischaracteristicofvonWillebrand"sdisease.
Inadditiontoitsroleinplateletplugformation,vWFisalsoresponsibleforthebindingandtransportoffactorVIII(antihemophilicfactor)inplasma(15).ItappearsthatthislattereventisresponsibleforboththestABIlityandeffectivedeliveryoffunctionalfactorVIII.StudiesindicatethatfactorVIIIbindstotheNH2-terminalportionofthematurevWFsubunitwithastoichiometryofonefactorVIIImoleculepervWFmonomer(16,17).
ThesinglechainvWFmonomercontainsalargenumberofcysteineresiduesatboththeNH2-terminalandCOOH-terminalends,whichareinvolvedinthemultimerformation.Carbohydrateanalysesindicatethatnearly15%ofthemassofvWFiscontributedbycarbohydrate(18).ItappearsthatthecarbohydrateservestoprotectvWFfromproteolysis,butisnotnecessaryforfunctionalactivityormultimerformation.
vWFispreparedfromcitratedhumanplasmausingacombinationoftheproceduresdescribedbyThorell(19),andLollar(20).AfactorVIII"free"vWFpreparation,furtherpurifiedtoensureremovaloffactorVIIIprocoagulantactivity(antigenstillpresent),isalsoavailable.Thepreparationsare>95%pureasjudgedbySDS-PAGEunderreducingconditions,andconsistoflargemultimersasdeterminedbyelectrophoresisinSDS/agarosegels.Theproteinisshippedfrozenin0.025Msodiumcitrate,0.1Mglycine,0.1MNaCl,pH6.8,forstorageat-70°C.
Properties:
| Localization/th> | Plasmaandsubendothelium |
|---|---|
| Modeofaction | facilitatesplateletplugformationbyformingabridgebetweenplateletglycoproteinIBandexposedcollageninthesubendothelium;alsobindsandtransportsfactorVIII |
| Molecularweight | 260,000to>10,000,000(1-8) |
| Isoelectricpoint | 5.7-5.9(21) |
| Extinctioncoefficient | NotApplicable;concentrationdeterminedbytotalproteinassay. |
| Concentrationinplasma | 10micrograms/mL(21) |
| Structure | multimericproteincomposedofidentical260,000molecularweightsubunits |
| Percentcarbohydrate | approximately15%(18) |
