| Formulation | 50%glycerol/water(v/v) |
| Storage | -20°C |
| Purity | >95%bySDS-PAGE |
| ActivityDetermination | Clottingassay |
| ShelfLife(properlystored) | 6months |
TheactivationofsinglechainfactorV(330,000)byalpha-thrombinisrepresented.ArrowsindicatethesiteatwhichfactorViscleavedbythrombintoyieldtwoactivationpeptides(71,000and120,000),inadditiontotheheavy(94,000)andlight(74,000)chainswhichformVa.TheheavyandlightchainsoffactorVaremainnon-covalentlyassociatedinthepresenceofcalciumions.
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanFactorV,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
| SpecialNotes | FactorVisaverylABIleprotein.Somedegradedfragmentsareusuallynoticeableinpreparations. |
Overview:
FactorVisalarge,singlechain,plasmaglycoproteinwhichisanessentialcomponentinthebloodcoagulationcascade(1).Duringcoagulation,theprocofactor,factorV,isconvertedtotheactivecofactor,factorVa,vialimitedproteolysisbytheserineproteasealpha-thrombin(illustratedabove),andlessefficientlybyfactorXa.TheactivecofactoriscomposedofanNH2-terminalderivedheavychain(Mr=94,000)andaCOOH-terminalderivedlightchain(Mr=74,000)whichremainnon-covalentlyassociatedinthepresenceofcalciumions.FactorVaservesasacofactorfortheserineproteasefactorXa.FactorVaandXaassembleonaphospholipidsurfaceinanon-covalentandcalciumion-dependentmanner,toformtheprothrombinasecomplex.TheprothrombinasecomplexisresponsIBLefortherapidconversionofthezymogenprothrombintotheactiveserineprotease,alpha-thrombin.Assemblyoftheprothrombinasecomplexincreasestherateatwhichprothrombinisconvertedtothrombinbynearly300,000-foldrelativetotheratewithfactorXaalone.DownregulationoftheprothrombinasecomplexisaccomplishedpartlythroughtheinactivationoffactorVabyactivatedproteinC.
HumanfactorVispreparedfromfreshfrozenhumanplasmausingimmunoaffinitychromatographyasdescribedbyKatzmannandcoworkers(2).BovinefactorVispreparedfromfreshbovineplasmausingconventionalchromatographictechniquesasdescribedbyNesheimandcoworkers(3).PurifiedfactorVissuppliedin50%glycerol(vol/vol),andshouldbestoredat-20°C.PurityisdeterminedbySDS-PAGEanalysisandactivityismeasuredinafactorVclottingassay.
Properties:
| Localization | Plasmaandplatelets | |||||
|---|---|---|---|---|---|---|
| Plasmaconcentration | 7.0µg/ml(human)(4,5) 35µg/ml(bovine)(5) | |||||
| Modeofaction | Procofactor;activatedbythrombintoformtheactivecofactor,factorVa | |||||
| Molecularweight | 330,000(3)-(determinedforbovinefactorV) | |||||
| Extinctioncoefficient |
| |||||
| Structure | onesubunit,2196aminoacids(1,6)-(determinedforhumanfactorV) | |||||
| Percentcarbohydrate | approximately25%(baseduponthecalculatedmolecularweightofhumanfactorV) |
