Formulation50%(vol/vol)Glycerol/H2OMolecularWeightStorage-20°CPurity>95%bySDS-PAGECompoundAssayThrombininhibitionShelfLife(properlystored)12monthsChemicalFormulaInhibitionofThrombinbyHeparinCofactorIIThrombin(IIa),heparin(HEP)andheparincofactorII(HCII)interacttoformaternarycomplexviaarandomorderbireactantmechanism.HeparinsubsequentlydissociatesfromthecomplexasthrombinformsacovalentcomplexwiththeheparincofactorII.SampleGelInformation:GelNovex4-12%Bis-TrisLoadHumanHeparinCofactorII,1µgperlaneBufferMOPSStandardSeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa)Overview:HeparincofactorII(HCII)(heparincofactorA,antithrombinBM,dermatansulfatecofactor,humanleuserpin-2)isasinglechainglycoprotein(Mr=65,600,10%carbohydrate)memberoftheserineproteaseinhibitor(serpin)family(1-3).AminoacidsequenceanalysisofheparincofactorIIreveals3potentialN-glycosylationsitesand2sulfatedtyrosineslocatedintwointernalrepeatsof7residueslocatedneartheNH2-terminal(4-6).Theplasmaconcentrationisestimatedtobe90µg/ml(2).Inthecoagulationcascade,heparincofactorIIinhibitsthrombinbyformationofabimolecularcomplexinthepresence(k2=4.5x108M-1min-1)andabsence(k2=5.0x105M-1min-1)ofheparin(2,7-9).ThiscomplexisstabletodenaturantsandessentiallyirreversIBLe.Inadditiontoheparin,awidevarietyofpolyanioniccompoundsaccelerateinhibitionofthrombinbyHCII(3).Noteworthyamongthesecompoundsisdermatansulfate,whichacceleratesinhibitionofthrombinbyHCII,butnotantithrombinIII(1,6)ThisfeaturehasbeenutilizedtodevelopaplasmabasedassayspecificforheparincofactorII.AlthoughHCIIshowsonly~25%sequencehomologytootherserpins,thereactivesitepeptide(COOH-terminal36aminoacids)releasedoninhibitionofthrombinis53%homologoustothatreleasedfromantithrombinIII(ATIII)bythrombin(4,9).ThescissilebondinHCIIisLeu-Serwhichisunusualforanargininespecificproteaseinhibitor(4,9).IncontrasttoATIII,theonlycoagulationproteaseinhibitedbyHCIIisthrombin,OtherproteasesinhibitedbyHCIIincludeα-chymotrypsin,neutrophilcathepsinGandstreptomycesgriseusproteaseB(3).ThemajorphysiologicroleofheparincofactorIImayremaintobediscovered.HeparincofactorIIispreparedfromfreshfrozenplasmabyamodificationoftheprocedureofGriffith,etal.,(9).PurityisassessedonSDSPAGEandactivityisdeterminedbasedonthrombininhibitioninthepresenceofdermatansulfate.HCIIissuppliedin50%(vol/vol)glycerol/H20forstorageat-20°C.Properties:LocalizationPlasmaPlasmaconcentration90µg/ml(2)Modeofactionserineproteaseinhibitor;inhibitsthrombin,a-chymotrypsin,neutrophilcathepsinG,streptomycesgriseusproteaseBMolecularweight65,600(2)ExtinctioncoefficientE1%1cm,280nm=5.93(5)Isoelectricpoint4.95-5.15(2)Structuresinglechainglycoprotein,3potentialN-glycosylationsites,two7-aminoacidresiduerepeats,reactivesitesequence:TVTTVGFMPL-STQVRFTVDR(4)Percentcarbohydrate9.9%(2)Post-translationalmodifications3sulfatedtyrosines