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1、J.gen.Virol.(1985),66,363-366.PrintedinGreatBritain363Keywords:coronavirus/leupeptin/proteaseinhibitorInhibitionoftheGrowthofHumanCoronavirus229EbyLeupeptinByGEOFFREYAPPLEYARD*ANDMARGARETTISDALEWellcomeResearchLaboratories,LangleyCourt,Beckenham,KentBR33BS,U.K.(Ac
2、cepted17October1984)SUMMARYTheproteaseinhibitorleupeptinpreventedmultiplicationofthehumancoronavirusstrain229EinculturesofMRC-Ccells.TheICsoofleupeptininplaquereductiontestswas0.4~tg/ml,whereasgrowthofhostcellswasunaffectedbyleupeptinat50~tg/ml.Inhibitionofplaquef
3、ormationcouldbepreventedbytheadditionofproteasestotheoverlaymedium.Insingle-cyclegrowthexperiments,leupeptinreducedvirusyieldonlyifaddedwithin2hofinfection,indicatingitsactiononanearlystageofvirusreplication.Proteolyticprocessingofviralproteinsisanessentialstagein
4、thegrowthofvirusesbelongingtomanydifferentfamilies.Althoughthereisnodirectevidencefortheinvolvementofproteasesinthereplicationofcoronaviruses,recentreportshavedescribedtheabilityoftrypsintostimulatethegrowthofsomecoronavirusesincellculture.Forexample,trypsinfacili
5、tatesplaqueformationbynumerousstrainsofavianinfectiousbronchitisvirusinchickembryocellcultures(Otsuki&Tsubokura,1981),byanenteropathogenicbovinecoronavirusinfoetalbovinethyroidorbraincells(Storzetal.,1981)andbyvariantsofmousehepatitisvirusinculturesofSRCDF1DBTmous
6、ecells(Yoshikura&Tejima,1981).Trypsinalsogreatlyincreasestheyieldofneonatalcalfdiarrhoeacoronavirusfrombovineembryoniclungcellsunderliquidmedium(Toth,1982).Thesefindingssuggesttheinvolvementofaprotease,possiblyoccurringinsuboptimalamountsinsometypesofcell,inthegro
7、wthofavarietyofcoronaviruses.Theydonot,however,identifythestage(s)ofvirusreplicationthatmaybeaffected.Wehavenowobservedthattheproteaseinhibitor,leupeptin,suppressesreplicationofthehumanrespiratorycoronavirus229EinculturesofMRC-Ccells.Theeffectappearstobeexertedear
8、lyinthevirusgrowthcycle.TheMRC-Ccellline,derivedoriginallyfromMRC5cells,wasobtainedfromDrA.F.Bradburne.Humancoronavirusstrain229E(HCV229E),providedbyDrS
