Formulation	50%(vol/vol)Glycerol/H2O
MolecularWeight
Storage	-20°C
Purity	>95%bySDS-PAGE
Compound
Assay	Thrombininhibition
ShelfLife(properlystored)	12months
ChemicalFormula

DomainStructureofAntithrombinIII
AntithrombinIIIcontainsthreeintra-chaindisulfidebonds(-S-S-),acarbohydraterichdomain(CHO),anNH2-terminalheparinbindingdomain,andaCOOH-terminalserineproteasebindingdomain.

SampleGelInformation:

Gel	Novex4-12%Bis-Tris
Load	HumanATIII,1µgperlane
Buffer	MOPS
Standard	SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa)

Overview:

AntithrombinIII(ATIII)isasinglechainglycoproteinwithamolecularweightof58,000.Itisamemberoftheserpin(serineproteaseinhibitor)superfamilyandisconsideredtobethemostimportantinhibitorinthecoagulationcascade(1,2).ATIIIinhibitsawidespectrumofserineproteasesincludingthrombin,factorsIXa,XaandXIa,kallikrein,plasmin,urokinase,C1-esterase,andtrypsin.Themechanismofinhibitioninvolvestheformationofastable1:1complexbetweentheactivesiteoftheproteaseandthescissilebond(Arg385-Ser386)ofATIII.TheactivesiteserineofthrombinhasbeenshowntoformacovalentintermediatewiththeP1aminoacid(Arg385)ofATIII.TherateofinhibitionofserineproteasesbyATIIIisincreasedtovaryingdegreesbyheparin.InthecaseofbeingathrombininhibitororfactorXainhibitor,theinteractionwithATIIIisenhanced3ordersofmagnitudeinthepresenceofheparin.TheinteractionbetweenATIIIandheparininvolvesauniquesequenceofsulfatedandnon-sulfatedmonosaccharideunitsonheparin,andcriticallysineresiduesonATIII.ThebindingofATIIItoheparinoidstructuresonvascularendotheliumhasbeendemonstratedandshowntoenhancetheinhibitionoffactorsIXa,Xa,andthrombin.

ATIIImayalsofunctioninthecomplementcascade.ThebindingofATIIItofluidphasecomplementattack-complexesinserahasbeendemonstrated.Inaddition,theSproteinofcomplement(aninhibitorofthemembraneattack-complex)interfereswiththeATIII/thrombininteraction.

ATIIIispreparedfromfreshfrozenplasmabyheparin-agaroseaffinitychromatography(3).Thepurifiedproteinissuppliedin50%(vol/vol)glycerol/H2Oandshouldbestoredat-20°C.PurityisdeterminedbySDS-PAGEanalysis.

Properties:

Localization	Plasma
PlasmaConcentration	150µg/ml
Modeofaction	Serineproteaseinhibitor
Molecularweight	58,000(3)
Extinctioncoefficient	E 1% 1cm,280nm =6.2(4)
IsoelectricPoint	4.9-5.3(5)
Structure	Singlechain,threeintrachaindisulfidebonds(Cys8-Cys128,Cys21-Cys95,Cys239-Cys422)(4),10%a-helix,30-40%b-structure,50%randomcoil(5),scissilebond(Arg385-Ser386)
Percentcarbohydrate	9%(5,7)

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