Expression, purification and sublocalization of SARS-CoV nucleocapsid protein in insect cells.
Identifieur interne : 002A74 ( PubMed/Corpus ); précédent : 002A73; suivant : 002A75Expression, purification and sublocalization of SARS-CoV nucleocapsid protein in insect cells.
Auteurs : Ai-Xia Ren ; You-Hua Xie ; Yu-Ying Kong ; Guan-Zhen Yang ; Yao-Zhou Zhang ; Yuan Wang ; Xiang-Fu WuSource :
- Acta biochimica et biophysica Sinica [ 1672-9145 ] ; 2004.
English descriptors
- KwdEn :
- Animals, Blotting, Western, Cell Line, Cytoplasm (chemistry), Electrophoresis, Polyacrylamide Gel, Escherichia coli (metabolism), Humans, Insecta, Microscopy, Confocal, Microscopy, Fluorescence, Nucleocapsid Proteins (chemistry), Nucleocapsid Proteins (isolation & purification), Nucleocapsid Proteins (metabolism), Plasmids (metabolism), Protein Conformation, Recombinant Proteins (chemistry), Recombinant Proteins (metabolism), SARS Virus (metabolism).
- MESH :
- chemical , chemistry : Nucleocapsid Proteins, Recombinant Proteins.
- chemistry : Cytoplasm.
- chemical , isolation & purification : Nucleocapsid Proteins.
- metabolism : Escherichia coli, Nucleocapsid Proteins, Plasmids, Recombinant Proteins, SARS Virus.
- Animals, Blotting, Western, Cell Line, Electrophoresis, Polyacrylamide Gel, Humans, Insecta, Microscopy, Confocal, Microscopy, Fluorescence, Protein Conformation.
Abstract
The causative agent of severe acute respiratory syndrome (SARS) is a previously unidentified coronavirus, SARS-CoV. The nucleocapsid (N) protein of SARS-CoV is a major viral protein recognized by acute and early convalescent sera from SARS patients. To facilitate the studies on the function and structure of the N protein, this report describe the expression and purification of recombinant SARS-CoV N protein using the baculovirus expression system. Recombinant hexa-histidine-tagged N protein with a molecular mass of 47 kD was produced in insect cells. Recombinant N protein was purified to near homogeneity by Ni2+-NTA affinity chromatography. In addition, we examined the subcellular localization of the N protein by confocal microscopy in Trichoplusia ni BT1 Tn 5B1-4 cells infected with recombinant baculovirus. The N protein was found localized in the cytoplasm as well as in the nucleolus. The purified recombinant N protein can be used in further functional study of SARS-CoV.
DOI: 10.1093/abbs/36.11.754
PubMed: 15514849
Links to Exploration step
pubmed:15514849Le document en format XML
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<author><name sortKey="Ren, Ai Xia" sort="Ren, Ai Xia" uniqKey="Ren A" first="Ai-Xia" last="Ren">Ai-Xia Ren</name>
<affiliation><nlm:affiliation>Institute of Biochemistry and Biotechnology, College of Life Science, Zhejiang University, Hangzhou 310029, China.</nlm:affiliation>
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<author><name sortKey="Xie, You Hua" sort="Xie, You Hua" uniqKey="Xie Y" first="You-Hua" last="Xie">You-Hua Xie</name>
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<author><name sortKey="Kong, Yu Ying" sort="Kong, Yu Ying" uniqKey="Kong Y" first="Yu-Ying" last="Kong">Yu-Ying Kong</name>
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<author><name sortKey="Yang, Guan Zhen" sort="Yang, Guan Zhen" uniqKey="Yang G" first="Guan-Zhen" last="Yang">Guan-Zhen Yang</name>
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<author><name sortKey="Zhang, Yao Zhou" sort="Zhang, Yao Zhou" uniqKey="Zhang Y" first="Yao-Zhou" last="Zhang">Yao-Zhou Zhang</name>
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<author><name sortKey="Wang, Yuan" sort="Wang, Yuan" uniqKey="Wang Y" first="Yuan" last="Wang">Yuan Wang</name>
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<author><name sortKey="Wu, Xiang Fu" sort="Wu, Xiang Fu" uniqKey="Wu X" first="Xiang-Fu" last="Wu">Xiang-Fu Wu</name>
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<sourceDesc><biblStruct><analytic><title xml:lang="en">Expression, purification and sublocalization of SARS-CoV nucleocapsid protein in insect cells.</title>
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<affiliation><nlm:affiliation>Institute of Biochemistry and Biotechnology, College of Life Science, Zhejiang University, Hangzhou 310029, China.</nlm:affiliation>
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<author><name sortKey="Kong, Yu Ying" sort="Kong, Yu Ying" uniqKey="Kong Y" first="Yu-Ying" last="Kong">Yu-Ying Kong</name>
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<author><name sortKey="Yang, Guan Zhen" sort="Yang, Guan Zhen" uniqKey="Yang G" first="Guan-Zhen" last="Yang">Guan-Zhen Yang</name>
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<term>Electrophoresis, Polyacrylamide Gel</term>
<term>Escherichia coli (metabolism)</term>
<term>Humans</term>
<term>Insecta</term>
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<term>Microscopy, Fluorescence</term>
<term>Nucleocapsid Proteins (chemistry)</term>
<term>Nucleocapsid Proteins (isolation & purification)</term>
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<term>Blotting, Western</term>
<term>Cell Line</term>
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<front><div type="abstract" xml:lang="en">The causative agent of severe acute respiratory syndrome (SARS) is a previously unidentified coronavirus, SARS-CoV. The nucleocapsid (N) protein of SARS-CoV is a major viral protein recognized by acute and early convalescent sera from SARS patients. To facilitate the studies on the function and structure of the N protein, this report describe the expression and purification of recombinant SARS-CoV N protein using the baculovirus expression system. Recombinant hexa-histidine-tagged N protein with a molecular mass of 47 kD was produced in insect cells. Recombinant N protein was purified to near homogeneity by Ni2+-NTA affinity chromatography. In addition, we examined the subcellular localization of the N protein by confocal microscopy in Trichoplusia ni BT1 Tn 5B1-4 cells infected with recombinant baculovirus. The N protein was found localized in the cytoplasm as well as in the nucleolus. The purified recombinant N protein can be used in further functional study of SARS-CoV.</div>
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<Title>Acta biochimica et biophysica Sinica</Title>
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<Abstract><AbstractText>The causative agent of severe acute respiratory syndrome (SARS) is a previously unidentified coronavirus, SARS-CoV. The nucleocapsid (N) protein of SARS-CoV is a major viral protein recognized by acute and early convalescent sera from SARS patients. To facilitate the studies on the function and structure of the N protein, this report describe the expression and purification of recombinant SARS-CoV N protein using the baculovirus expression system. Recombinant hexa-histidine-tagged N protein with a molecular mass of 47 kD was produced in insect cells. Recombinant N protein was purified to near homogeneity by Ni2+-NTA affinity chromatography. In addition, we examined the subcellular localization of the N protein by confocal microscopy in Trichoplusia ni BT1 Tn 5B1-4 cells infected with recombinant baculovirus. The N protein was found localized in the cytoplasm as well as in the nucleolus. The purified recombinant N protein can be used in further functional study of SARS-CoV.</AbstractText>
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