GuanylinPrecursor,Guanylatecyclaseactivator2A,Guanylatecyclase-activatingprotein1,Gap-IGUCA2,STARA,GUANYLIN.
Heat-stableenterotoxins(STa)aresmall,cysteine-richpeptidessecretedbyEscherichiacolithatareabletoinducediarrheathroughthestimulationofanintestine-specificreceptor-guanylylcyclaseknownasSTaR.BindingofSTatoSTaRinducesadramaticincreaseinthecGMPcontentofthecell;theincrease,inturn,inhibitssaltabsorptionandstimulateschloridesecretion.Thisimbalanceofionsisaccompaniedbyamassiveaccumulationofwaterinthegutthatgivesrisetothediarrheaanddehydrationcharacteristicofenterotoxinactivity.TheidentificationofareceptorforSTaonintestinalbrushbordermembranessuggestedtheexistenceofanendogenousactivator,describedguanylin,a15-aminoacidpeptidepurifiedfromratsmallintestine,asapotentialligandfortheSTaR.ThispeptidesharessequencesimilaritywithSTa;seealsouroguanylin.Themolecularcloningofthehumanandmouse
CDNAsencodingguanylinwasreported.ThesequencesdemonstratedthatguanylinispresentattheC-terminalendofalargerprecursorprotein.Expressioninmammaliancellsindicatedthatthe94-aminoacidproguanylinisinactive.The
BIOLOGicallyactiveguanylincanbereleasedbyeitherchemicalorenzymatictreatmentofproguanylin.ByNorthernblotanalysisandinsituhybridization,showedthatexpressionofguanylinmRNAisrestrictedtocellsoftheintestinalepithelium,specificallythePanethcellsatthebaseofthesmallintestinalcrypts.TheseresultsdemonstratethatguanylinisanendogenousactivatorofSTaRisolatedacDNAencodinganapparentprecursorofguanylinfromahumanintestinalcDNAlibrary.ThemRNAwasexpressedathighlevelsinhumanileumandcolon.Inthemouse,interspecificbackcrossanalysisusedtomaptheGuca2genetothedistalhalfofmousechromosome4inaregionofhomologywithhumanchromosome1p.ByfluorescenceinsituhybridizationmappedtheGUCA2genetohuman1p35-p34Guanylinisthoughttomodulateintestinalwater/electrolytetransportinaparacrinemodereportedthenucleotidesequenceofthegene,thecharacteristicsofitscirculatingmolecularform,anditslocalizationinenterochromaffincellsofthegut.Thegene,approximately2.6kbinsize,consistsof3exonsinterruptedby2introns.Thehormonalformofguanylinisa94-aminoacidpeptidewithamolecularmassof10.3kDa.Guanylinissynthesizedbygutenterochromaffincellsasaprohormoneof115aminoacidsandisprocessedtothemolecularformof94aminoacidscirculatingintheblood.
ProguanylinHumanRecombinantproducedinE.coliisasingle,non-glycosylatedpolypeptidechain(a.a22-115)containing104aminoacidsincludinga10a.aN-terminalHistag.Thetotalmolecularmassis11.5kDa(calculated).
EscherichiaColi.
SterileFilteredWhitelyophilized(freeze-dried)powder.
Proguanylinfiltered(0.4µm)andlyophilizedin0.5mg/mlindeionizedH20.
Itisrecommendedtoadddeionizedwatertoprepareaworkingstocksolutionofapproximately0.5mg/mlandletthelyophilizedpelletdissolvecompletely.Proguanylinisnotsterile!Pleasefiltertheproductbyanappropriatesterilefilterbeforeusingitinthecellculture.
Storelyophilizedproteinat-20°C.Aliquottheproductafterreconstitutiontoavoidrepeatedfreezing/thawingcycles.Reconstitutedproteincanbestoredat4°Cforalimitedperiodoftime;itdoesnotshowanychangeafteroneweekat4°C.
Greaterthan90.0%asdeterminedbySDS-PAGE.
MKHHHHHHASVTVQDGNFSFSLESVKKLKDLQEPQEPRVGKLRNFAPIPGEPVVPILCSNPNFPEELKPLCKEPNAQEILQRLEEIAEDPGTCEICAYAACTGC.
Prospec"sproductsarefurnishedforLABORATORYRESEARCHUSEONLY.Theproductmaynotbeusedasdrugs,agriculturalorpesticidalproducts,foodadditivesorhouseholdchemicals.