| Formulation | 50%glycerol/water(v/v) |
| Storage | -20°C |
| Purity | >95%bySDS-PAGE |
| ActivityDetermination | Clottingassay |
| ShelfLife(properlystored) | 12months |
ThedomainstructureoffactorXisrepresented,where:GLA=regioncontainingγ-carboxyglutamicacidresidues,EGF=regioncontainingsequenceshomologoustohumanepidermalgrowthfactor,AP=activationpeptidereleaseduponconversionofthezymogentotheactiveserineprotease,CATALYTICDOMAIN=regioncontainingtheserineproteasecatalytictriad.ThearrowindicatesthesitewhichisproteolyticallycleavedbyfactorXaseduringactivationofthezymogen.
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanFactorX,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
Overview:
FactorXisavitaminK-dependentproteinzymogenwhichissynthesizedintheliverandcirculatesinplasmaasatwochainmoleculelinkedbyadisulfidebond(1,2).Priortosecretionintoplasma,post-translationalmodificationsproduce11gamma-carboxyglutamicacid(gla)residuesandasingleb-hydroxyasparticacidresidue,whicharelocatedwithintheNH2-terminallightchain.Thelightchainalsocontainstwoepidermalgrowthfactor(EGF)homologydomains.TheCOOH-terminalheavychainoffactorXcontainsmostofthecarbohydratemoieties,aswellasthelatentserineproteasedomain.TheactivationoffactorXiscatalyzedbyeithertheintrinsicfactorXasecomplex(factorIXa,factorVIIIa,cellularsurfaceandcalciumions)ortheextrinsicfactorXasecomplex(factorVIIa,tissuefactor,cellularsurfaceandcalciumions).ActivationofhumanfactorXbyeithercomplexresultsincleavageatArg52-Ile53oftheCOOH-terminalheavychainandsubsequentreleaseofa52aminoacidactivationglycopeptide.FactorXathenservesastheenzymecomponentoftheprothrombinasecomplexwhichisresponsIBLefortherapidconversionofprothrombintothrombin.TheglaresiduesenablefactorX/Xatobindphospholipid(i.e.cellsurfaces)inacalciumdependentmanner;arequirementforassemblyoftheprothrombinasecomplex.ThefirstEGFhomologydomaincontainsaCa2+bindingsitewhichactsasahingetofoldtheEGFandGLAdomainstowardseachother(12).Thisregionofthemoleculeisinvolvedintherecognitionofcellularbindingdomains.
HumanfactorXisisolatedfromfreshfrozenhumanplasmabyacombinationofconventionaltechniques(3)andimmunoaffinitychromatography(4).InadditiontothestandardhumanfactorXpreparation,Gla-domainlesshumanfactorXisalsoavailable.BovinefactorXisisolatedfromfreshbovineplasmausingamodificationoftheprocedurereportedbyBajajetal.(5,6).Thepurifiedzymogenissuppliedin50%(vol/vol)glycerol/H2Oandshouldbestoredat-20oC.PurityisdeterminedbySDS-PAGEanalysisandactivityismeasuredinafactorXclottingassay.
Properties:
| Localization | Plasma | ||||||||
|---|---|---|---|---|---|---|---|---|---|
| Plasmaconcentration | 10µg/ml | ||||||||
| Modeofaction | Zymogen;precursortotheserineproteasefactorXa | ||||||||
| Molecularweight | 58,900(human)(7) 55,100(bovine)(8) | ||||||||
| Extinctioncoefficient |
| ||||||||
| Isoelectricpoint | 4.9-5.2(human)(9) 4.8-5.2(bovine)(9) | ||||||||
| Structure | twosubunits,Mr=16,200and42,000(human),Mr=16,500and39,300(bovine),NH2-terminalgladomain,andtwoEGFdomains | ||||||||
| Percentcarbohydrate | 15%(human)(7) 10%(bovine)(8) | ||||||||
| Post-translationalmodifications | elevenglaresidues(7,8) oneβ-hydroxyaspartate |
