| Formulation | 50mMNaPO4,150mMNaCl,pH7.4 |
| Storage | -80°C |
| Purity | >95%bySDS-PAGE |
| ActivityDetermination | N/A |
| ShelfLife(properlystored) | 12months |
Thedomainstructureofhumanvitronectinisrepresented,where:RGDsequencerepresentsthemajorcellattachmentsite,anacidic(---)regionneartheNH2-terminusrepresentsanumberofmacromolecularbindingsites,theshadedregionrepresentsthehemopexin-typedomain,thebasic(+++)regionneartheCOOH-terminusrepresentstheheparin-bindingsite,thearrowrepresentsanendogenousproteasecleavagesite,PO4isthesiterepresentingphosphorylationduetoproteinkinaseA.
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanVitronectin,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
Overview:
Vitronectinisaplasmaglycoproteinthatcirculatesinthebloodataconcentrationrangeof200to400mg/mlconstituting0.2to0.5%oftotalplasmaproteins(1,2).Itisdetectedasamixtureofboth75kDaand65kDaforms(3).The65kDaformisaproductofproteolyticprocessingoftheformerwhichappearstobeendogenouslypresent.Vitronectinisinvolvedinanumberofphysiologicalprocessesthatincludebloodcoagulation,fibrinolysis,complementcascade,andcelladhesion.Vitronectin(SerumspreADIngfactor)isoneofthetwomajorcell-adhesiveglycoproteins.Itisamajorligandforthevitronectinreceptor(αV/β3)orαV/β5integrinonadhesivecells(4).Otherfunctionsincludeheparin-binding;collagen-binding;osteonectin-binding;complementlysisinhibitorand“scavenger”proteinformacromolecularproductsofthecomplementandhaemostasiscascadesystems(5).Italsofunctionsasasubstratefortransglutaminase(FactorXIIIa)crosslinking,cAMP-dependentproteinkinase,andtyrosylproteinsulfotransferase(4,5).Vitronectinhasimplicationsforthrombosisduetoitsheparinbindingproperties.IthasbeenpostulatedthatvitronectinpreventstheaccelerationofclotformationinhibitionbyactingasaheparinscavengerultimatelyprotectingthrombinandfactorXafromheparin-dependantinactivationbyantithrombinIIIorheparincofactorII(6,7).VitronectinisalsothoughttoplayanimportantroleinfibrinolysisbyitsABIlitytobindactivePAI-1(4).Extracellularbindingofvitronectintoplasminogenactivatorinhibitor-1(PAI-1)andplasminogen,stabilizetheinhibitorandthusaffecttissueplasminogenactivator-mediatedplasminformation(4).
ThevitronectindomainstructurestartingattheNH4-terminusconsistsofasomatomedin-Bregion(residues1to44),aconnectingsegmentcontainingthemajorcellattachmentRGDsequence(residues45to47)andahighlyacidicregion(residues53to64),followedbyhemopexin-likerepeats(homologytohemopexinwhichisaheme-bindingplasmaprotein),aglycosaminoglycan-bindingsitewhichisrepresentedbya40-aminoacidstretchrichinbasicresidues,aprotease-sensitiveregionsusceptIBLetohighconcentrationsofthrombin,andaCOOH-terminalproteinkinaseA-dependentphosphorylationsite,whichisadjacenttotheproteasecleavagesite(4,5).
HumanvitronectinispreparedfromfreshfrozenplasmasimilartotheproceduredescribedbyYatohgoandcoworkers(8).Purifiedvitronectinissuppliedin50mMsodiumphosphate,150mMNaCl,pH7.4andshouldbestoredat-80°C.PurityisassessedbySDS-PAGEanalysis.
Properties:
| Localization | Plasma |
|---|---|
| ModeofAction | Vitronectinprovidesamolecularlinkimportantforcelladhesionandregulationofdefensemechanismsintheterminalstepsofbloodcoagulationandcomplementcascades |
| PlasmaConcentration | 200-400mg/ml(1,2) |
| MolecularWeight | 75,000single-chainformandatwo-chainformcomposedof10,000and65,000(SDS-PAGE)(4) |
| Extinctioncoefficient | E1%1cm,280nm=13.8(9) |
| IsoelectricPoint | 4.75–5.25(8Murea)(4) |
| Structure | Circulatesinmonomeric,dimericandpossiblyhigheroligomericforms.Monomer:459aminoacid,singlechainpolypeptidewithsevenintrachaindisulfidesandonefreesulfhydryl |
| CarbohydrateContent | 10-15%(w/w) |
