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1、VIROLOGY182,765-773(1991)IntracellularProcessingofthePorcineCoronavirusTransmissibleGastroenteritisVirusSpikeProteinExpressedbyRecombinantVacciniaVirusDAVIDJ.PULFORDANDPAULBRImONDivisionofMolecularBiology,A.F.R.C.,InstituteforAnimalHealth,ComptonLaboratory,
2、Compton,Newbury,Berkshire,RG16ONN,UnitedKingdomReceivedDecember18,1990;acceptedFebruary20,1991TheSpike(S)proteinfromavirulentBritishfieldisolateofporcinetransmissiblegastroenteritisvirus(TGEV)FS772/70wasconstructedfromcDNAandinsertedintothevacciniavirus(VV)
3、thymidinekinasegenelocusunderthecontroloftheVVearly/lategeneP7.5Kpromoter.RecombinantSproteinwassynthesizedasanendo-P-N-acetylglucosamini-daseH(EndoH)-sensitiveglycoproteinwithhighmannosesimpleoligosaccharides(gpl90)thatunderwentpost-trans-lationalmodificat
4、iontoanEndoH-resistantglycoproteinwithcomplexoligosaccharides(gp210).lmmunofluorescenceanalysisdemonstratedthatthemajorityofrecombinantSproteinwasretainedattheGolgibutsomeSproteinwasexpressedontheplasmamembrane.Monoclonalantibodies(mAbs)raisedagainstnativeS
5、proteinreactedwiththisrecombinantSprotein;also,miceinfectedwiththerecombinantvacciniavirus(rVV)expressingtheSproteininducedTGEVneutralizingantibodies.AtruncatedSprotein(SA)wasalsoexpressedinrVV-infectedcellsbyintroducingadeletionintotheSproteincDNAthatremov
6、ed292aminoacidsfromtheC-terminus.TheSAprotein(gpl70)wasshowntobeantigenicallysimilartoTGEVSproteinbyimmunofluorescenceandimmunoprecipitationtestsbutwasretainedintheendoplasmicreticulumandnotexpressedonthecellsurface.oISSIAcademicPWS.inc.INTRODUCTIONvirus(FI
7、PV),caninecoronavirus,andporcinerespira-torycoronavirus,arenotcleaved(forareviewseeTransmissiblegastroenteritisvirus(TGEV)belongsSpaanetal.,1988).However,theSproteinsfromtothefamilyCoronaviridae,alargegroupofenvelopedMHV,infectiousbronchitisvirus,bovinecoro
8、navirus,viruseswithapositive-strandedRNAgenome.Thehumancoronavirus(OC43and229E),andporcineviruscausesgastroenteritisinneonatalpigs,resultinghemagglutinatingencephalomyelitisviruswhichareinahighmortalit
