Antigenic characterization of severe acute respiratory syndrome-coronavirus nucleocapsid protein expressed in insect cells: The effect of phosphorylation on immunoreactivity and specificity.

Abstract

The nucleocapsid (N) protein of severe acute respiratory syndrome-coronavirus (SARS-CoV) is involved in the pathological reaction to SARS and is a key antigen for the development of a sensitive diagnostic assay. However, the antigenic properties of this N protein are largely unknown. To facilitate the studies on the function and antigenicity of the SARS-CoV N protein, 6x histidine-tagged recombinant SARS-CoV N (rSARS-N) with a molecular mass of 46 and 48kDa was successfully produced using the recombinant baculovirus system in insect cells. The rSARS-N expressed in insect cells (BrSARS-N) showed remarkably higher specificity and immunoreactivity than rSARS-N expressed in E. coli (ErSARS-N). Most of all, BrSARS-N proteins were expressed as a highly phosphorylated form with a molecular mass of 48kDa, but ErSARS-N was a nonphosphorylated protein. In further analysis to determine the correlation between the phosphorylation and the antigenicity of SARS-N protein, dephosphorylated SARS-N protein treated with protein phosphatase 1 (PP1) remarkably enhanced the cross-reactivity against SARS negative serum and considerably reduced immunoreactivity with SARS-N mAb. These results suggest that the phosphorylation plays an important role in the immunoreactivity and specificity of SARS-N protein. Therefore, the BrSARS-N protein may be useful for the development of highly sensitive and specific assays to determine SARS infection and for further research of SARS-N pathology.

• Authors: Gu-Choul Shin, Yoon-Seok Chung, In-Soo Kim, Hae-Wol Cho, Chun Kang
• Journal: Virus research (Virus Res) Vol. 127 Issue 1 Pg. 71-80 (Jul 2007) ISSN: 0168-1702 [Print] Netherlands
• PMID: 17499376 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antibodies, Viral
• Antigens, Viral
• Coronavirus Nucleocapsid Proteins
• Nucleocapsid Proteins

Topics

• Animals
• Antibodies, Viral (blood, immunology)
• Antibody Affinity
• Antibody Specificity (physiology)
• Antigens, Viral (immunology)
• Coronavirus Nucleocapsid Proteins
• Enzyme-Linked Immunosorbent Assay
• Insecta (cytology)
• Nucleocapsid Proteins (genetics, immunology, metabolism)
• Phosphorylation
• Severe acute respiratory syndrome-related coronavirus (genetics, immunology)