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ISSN 1672-9145                                                 Acta Biochim Biophys Sin 2004, 36(11): 754-758                                                   CN 31-1940/Q

Expression, Purification and Sublocalization of SARS-CoV Nucleocapsid Protein in Insect Cells

Ai-Xia REN1,2, You-Hua XIE2, Yu-Ying KONG2, Guan-Zhen YANG2, Yao-Zhou ZHANG1,3*, Yuan WANG2*, and Xiang-Fu WU2,3*

 

1Institute of Biochemistry and Biotechnology, College of Life Science, Zhejiang University, Hangzhou 310029, China;

2State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and cell Biology, Shanghai Institutes for Biological Sciences,

Chinese Academy of Sciences, Graduate School of the Chinese Academy of Sciences, Shanghai 200031, China;

3Institute of Biochemistry, Zhejiang University of Sciences, Hangzhou 310018, China

 

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.

 

Key words        severe acute respiratory syndrome (SARS); SARS coronavirus; nucleocapsid protein; baculovirus; insect cells

 

 

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Received: August 16, 2004        Accepted: October 8, 2004

*Corresponding author: Tel, 86-571-86843190; Fax, 86-571-86843198; E-mail, [email protected]