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2010-01-15Zeitschriftenartikel DOI: 10.1186/1743-422X-7-6
Human Coronavirus NL63 Open Reading Frame 3 encodes a virion-incorporated N-glycosylated membrane protein
dc.contributor.authorMüller, Marcel A.
dc.contributor.authorHoek, Lia van der
dc.contributor.authorVoss, Daniel
dc.contributor.authorBader, Oliver
dc.contributor.authorLehmann, Dörte
dc.contributor.authorSchulz, Axel R.
dc.contributor.authorKallies, Stephan
dc.contributor.authorSuliman, Tasnim
dc.contributor.authorFielding, Burtram C
dc.contributor.authorDrosten, Christian
dc.contributor.authorNiedrig, Matthias
dc.date.accessioned2018-05-07T13:44:00Z
dc.date.available2018-05-07T13:44:00Z
dc.date.created2010-02-26
dc.date.issued2010-01-15none
dc.identifier.otherhttp://edoc.rki.de/oa/articles/reXZoMxJ2MeM2/PDF/25ZYPlUUTJlSc.pdf
dc.identifier.urihttp://edoc.rki.de/176904/593
dc.description.abstractBackground: Human pathogenic coronavirus NL63 (hCoV-NL63) is a group 1 (alpha) coronavirus commonly associated with respiratory tract infections. In addition to known non-structural and structural proteins all coronaviruses have one or more accessory proteins whose functions are mostly unknown. Our study focuses on hCoV-NL63 open reading frame 3 (ORF 3) which is a highly conserved accessory protein among coronaviruses. Results: In-silico analysis of the 225 amino acid sequence of hCoV-NL63 ORF 3 predicted a triple membrane-spanning protein. Expression in infected CaCo-2 and LLC-MK2 cells was confirmed by immunofluorescence and Western blot analysis. The protein was detected within the endoplasmatic reticulum/Golgi intermediate compartment (ERGIC) where coronavirus assembly and budding takes place. Subcellular localization studies using recombinant ORF 3 protein transfected in Huh-7 cells revealed occurrence in ERGIC, Golgi- and lysosomal compartments. By fluorescence microscopy of differently tagged envelope (E), membrane (M) and nucleocapsid (N) proteins it was shown that ORF 3 protein colocalizes extensively with E and M within the ERGIC. Using N-terminally FLAG-tagged ORF 3 protein and an antiserum specific to the C-terminus we verified the proposed topology of an extracellular N-terminus and a cytosolic C-terminus. By in-vitro translation analysis and subsequent endoglycosidase H digestion we showed that ORF 3 protein is N-glycosylated at the N-terminus. Analysis of purified viral particles revealed that ORF 3 protein is incorporated into virions and is therefore an additional structural protein. Conclusions: This study is the first extensive expression analysis of a group 1 hCoV-ORF 3 protein. We give evidence that ORF 3 protein is a structural N-glycosylated and virion-incorporated protein.ger
dc.language.isoeng
dc.publisherRobert Koch-Institut
dc.subjectAnimalseng
dc.subjectWesterneng
dc.subjectMoleculareng
dc.subjectCell Lineeng
dc.subjectHumanseng
dc.subjectMicroscopyeng
dc.subjectModelseng
dc.subjectBiologicaleng
dc.subjectMacaca mulattaeng
dc.subjectBlottingeng
dc.subjectCoronavirus/geneticseng
dc.subjectEndoplasmic Reticulum/chemistryeng
dc.subjectGlycoproteins/geneticseng
dc.subjectGlycoproteins/metabolismeng
dc.subjectGolgi Apparatus/chemistryeng
dc.subjectLysosomes/chemistryeng
dc.subjectConfocaleng
dc.subjectFluorescenceeng
dc.subjectOpen Reading Frameseng
dc.subjectViral Matrix Proteins/geneticseng
dc.subjectViral Matrix Proteins/metabolismeng
dc.subjectViral Structural Proteins/analysiseng
dc.subjectVirion/chemistryeng
dc.subject.ddc610 Medizin
dc.titleHuman Coronavirus NL63 Open Reading Frame 3 encodes a virion-incorporated N-glycosylated membrane protein
dc.typeperiodicalPart
dc.identifier.urnurn:nbn:de:0257-1005923
dc.identifier.doi10.1186/1743-422X-7-6
dc.identifier.doihttp://dx.doi.org/10.25646/518
local.edoc.container-titleVirology Journal
local.edoc.fp-subtypeArtikel
local.edoc.type-nameZeitschriftenartikel
local.edoc.container-typeperiodical
local.edoc.container-type-nameZeitschrift
local.edoc.container-urlhttp://www.virologyj.com/content/7/1/6
local.edoc.container-publisher-nameBioMedCentral
local.edoc.container-volume7
local.edoc.container-issue6
local.edoc.container-year2010

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