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Title: Biochemical and functional characterization of proteins encoded by SARS-CoV subgenomic mRNA 8
Authors: LE TRA MY
Keywords: SARS-CoV, ORF 8a/8b, ORF 8ab, glycosylation, ubiquitination, IRF 3.
Issue Date: 15-Jun-2006
Citation: LE TRA MY (2006-06-15). Biochemical and functional characterization of proteins encoded by SARS-CoV subgenomic mRNA 8. ScholarBank@NUS Repository.
Abstract: The most striking difference in the subgenomic mRNA8 of SARS-CoV isolated from human and some animal species is the deletion of 29 nucleotides, resulting in splitting of a single ORF (ORF 8) into two ORFs (ORF 8a and 8b). ORF 8a and 8b are predicted to encode two small proteins, 8a and 8b, and ORF 8 a single protein 8ab (a fused form of 8a and 8b proteins). To understand the functions of these proteins, we cloned cDNA fragments covering these ORF into expression plasmids, and expressed these constructs in in vitro and in vivo systems. It was shown that expression of a construct containing ORF 8a and 8b generated only a single protein 8a; no detection of the 8b expression was obtained. Expression of a construct containing ORF 8 generated the 8ab fusion protein. Subcellular localization studies of HeLa cells expressing protein 8a, 8b and 8ab showed that 8a and 8ab are localized to the cytoplasm, and 8b in the ER. Site-directed mutagenesis and enzymatic treatment revealed that 8b and 8ab were modified by N-linked glycoprotein, and by ubiquitination. More interestingly, the two proteins could form complex with ubiquitin and interferon regulatory factor 3, suggesting the potential involvement of these protein in the pathogenesis of SARS-CoV.
Appears in Collections:Master's Theses (Open)

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