| Formulation | 50%glycerol/water(v/v) |
| Storage | -20°C |
| Purity | >95%bySDS-PAGE |
| ActivityDetermination | FactorVIIclottingassay |
| ShelfLife(properlystored) | 12months |
ThecomponentsoftheextrinsicfactorXaseenzymecomplexareillustrated.FactorVIIaanditscofactortissuefactor(TF)assembleonanegativelychargedmembranesurfaceinacalciumdependentmannertoformanenzymecomplexwhichproteolyticallyconvertsfactorXtofactorXa.ThecomplexisreferredtoastheextrinsicfactorXasecomplex,becausethetissuefactorcomponentispresumablyderivedfromasource"extrinsic"totheplasmaenvironment,whichisaccessIBLefollowingvascularinjury.
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanFactorVIIa,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
Overview:
HumanfactorVIIisasinglechainplasmaglycoprotein,andisazymogeninitsnativeform(1-4).ProteolyticactivationoffactorVII,yieldstheenzymefactorVIIa,whichwhenboundtotheintegralmembraneproteintissuefactor,formsanenzymecomplexthatproteolyticallyconvertsfactorXtofactorXa.ThisenzymecomplexisbestknownastheextrinsicfactorXase(pronouncedten-ase)complex,sincebyvirtueofitstissuefactorcomponent,itiscomposedofproteinnormallyextrinsictotheplasmaenvironment.
TheconversionoffactorVIItofactorVIIaiscatalyzedbyanumberofproteasesincludingthrombin,factorIXa,factorXa,factorXIa,andfactorXIIa.RapidactivationalsotakesplacewhenfactorVIIiscombinedwithtissuefactorinthepresenceofcalcium(5-9).Thislattereventisconsistentwithautocatalysis,andislikelyinitiatedbyasmallamountofpre-existingfactorVIIa.Theactivationreactionresultsincleavageofthepeptidebondbetweenarginine152andisoleucine153.
TheresultingfactorVIIaconsistsofanNH2-derivedlightchain(Mr=20,000),andaCOOH-terminalderivedheavychain(Mr=30,000),whichremainassociatedthroughasingledisulfidebond(cys135-cys262).Thelightchaincontainsthemembranebinding"Gladomain",whiletheheavychaincontainsthecatalyticdomain.
Unlikeotherserineproteases,factorVIIaaloneisnotreADIlyinhibitedbytheantithrombinIII/heparincomplex.However,inthepresenceoftissuefactor,antithrombinIII/heparinexhibitssignificantinhibitionoffactorVIIa(6).
HumanfactorVIIaispreparedfromaffinitypurifiedfactorVII,andpurifiedbyionexchangechromatography.Thepurifiedproteinissuppliedin50%(vol/vol)glycerol/H2Oandshouldbestoredat-20°C.ThepurityisdeterminedbySDS-PAGEanalysis,andactivityismeasuredinafactorVIIclottingassay.
Properties:
| Localization | Plasma | |||||
|---|---|---|---|---|---|---|
| Modeofaction | EnzymecomponentoftheextrinsicfactorXactivatingcomplex;alsoactivatesfactorIX,thusby-passingthecontactactivationsystem. | |||||
| Molecularweight | 50,000(human)(2) | |||||
| Extinctioncoefficient |
| |||||
| SpecificActivity | approximately16,000units/mg(determinedfromsinglestageclottingassay) | |||||
| Structure | 2subunits;NH2-terminalderivedlightchain(Mr=20,000),COOH-terminalderivedheavychain(Mr=30,000),NH2-terminalgla-domain,twoEGFdomains | |||||
| Percentcarbohydrate | 13%(10)-(baseduponanalysisofbovinefactorVII) | |||||
| Post-translationalmodifications | oneβ-hydroxyaspartate(11),tenglaresidues(12) |
