| Formulation | 20mMHepes,150mMNaCl,pH7.4 |
| MolecularWeight | |
| Storage | -80°C |
| Purity | >95%bySDS-PAGE |
| Compound | |
| Assay | N/A |
| ShelfLife(properlystored) | 12months |
| ChemicalFormula | |
Selectedproteolyticcleavagesiteswithinthehumanplasminogenmoleculeareillustrated.Cleavageofplasminogenbyelastaseyields"angiostatin"(kringles1-4)orangiostatin-likefragments(kringles1-3).HTI’s"angiostatin"productisderivedfromasinglecleavagewithintheplasminogenmoleculeresultinginaproductthatincludesKringles1-4aswellastheNH2-terminal78aminoacidsthatareabsentintheelastasegeneratedproduct.(K1-K5=the5kringledomains,B-CHAIN=catalyticdomainofplasmin,andthearrowsindicatethesitesofproteolyticcleavagebyplasmin,elastase,HTI’sproprietaryenzyme,andplasminogenactivators(PA"S)).
SampleGelInformation:
| Gel | Novex4-12%Bis-Tris |
|---|---|
| Load | HumanAngiostatin,1µgperlane |
| Buffer | MOPS |
| Standard | SeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa) |
Overview:
Angiostatinisasingle-chain,proteolyticfragmentofglu-plasminogenwhichhasamolecularweightofabout38,000.Itisapotentinhibitorofangiogenesisandwasfirstidentifiedandisolatedfromtheserumandurineoftumorbearingmice(1).Themouseproteinreportedlyextendsfromthreonine-98throughvaline-440(numberingfromtheNH2-terminusofglu-plasminogen).Thisfragmentofmouseplasminogen,aswellasthehumanequivalent,includesfouroutofthefivekringledomainsofplasminogen(1).
Theinhibitionofangiogenesisbyangiostatinisdirectlyrelatedtoinhibitingtheproliferationofendothelialcells(1,2).Becausetumorgrowthisknowntobeangiogenesisdependent,itwasinitiallyhypothesizedthattheinhibitorypropertiesofangiostatinwouldhaveclinicalutilityinarrestingvariouscancersthatareexpressedassolidtumors.Studiesperformedinanimalmodelsystemshavesincedemonstratedthatrecombinantangiostatineffectivelysuppressestumorgrowthandmetastasis(3,4).
Numerousenzymeshavebeenidentifiedwhichwillconvertplasminogentoangiostatinoratleasttoangiostatin-likefragments.Theenzymesincludeseveralmatrixmetalloproteinasesaswellasurokinaseandatumorcell-derivedreductase(5-10).ThepreciseenzymeormechanismwhichisresponsIBLefortheformationofangiostatininvivoisunknownanditisbelievedthattheremaybemultiplepathwaysfortheconversionofplasminogentoangiostatin.
Structure/functionstudieshaveindicatedthatthefirstthreekringledomains(andnotthefourth)areresponsiblefortheinhibitorypropertiesofangiostatin,andthatremovalofthefourthkringledomainmayactuallyyieldamorepotentinhibitor(11).
HTI’s"angiostatin"productisproducedbylimitedproteolysisofpurifiedhumanglu-plasminogenusingaproprietaryenzymepreparation.TheNH2-terminalsequenceofthisfragmentisidenticaltohumanglu-plasminogen(Glu-1)andthemoleculeterminatesatproline-452thusmakingitlargerthanthe"natural"orelastasederivedproductwithanapparentmolecularweightofabout50,000.Likeauthenticangiostatin,thisproductdemonstratesanantiproliferativeeffectwhentestedinagrowthfactor-inducedendothelialcellproliferationassay.Theproductisformulatedin20mMHepes,0.15MNaCl,pH7.4,andshouldbestoredfrozenat-70oCorbelow.
Properties:
| Modeofaction | Inhibitsendothelialcellproliferation | |||||
|---|---|---|---|---|---|---|
| Molecularweight | Approximately50,000bySDS-PAGE(appearsasadoubletduetothepresenceoftwocarbohydratevariants) | |||||
| Extinctioncoefficient |
| |||||
| Structure | Singlechain,4kringleregions,14disulfidebonds,nofreesulfhydryls | |||||
| Percentcarbohydrate | Approximately3% |
