Characterization of immunodominant linear B-cell epitopes on the carboxy terminus of the rinderpest virus nucleocapsid protein.
The nucleocapsid (N) protein of rinderpest virus (RPV) is one of the most abundant and immunogenic viral proteins expressed during natural or experimental infection. To identify immunogenic epitopes on the N protein, different forms of RPV N protein, including the full-length protein (N(1-525)), an amino-terminal construct (N(1-179)), and a carboxy-terminal construct (N(414-496)), were expressed in Escherichia coli as glutathione S-transferase (GST) fusion proteins. The antigenicity of each recombinant protein was evaluated by Western immunoblotting. All recombinants were recognized by hyperimmune RPV bovine antisera, indicating that immunoreactive epitopes may be present at both ends of the N protein. However, GST-N(414-496) was much more antigenic than GST-N(1-179) when tested with sera from vaccinated cattle, suggesting that an immunodominant or highly immunogenic epitope(s) may be located at the carboxy terminus of the N protein. Epitope mapping with overlapping peptides representing different regions of the carboxy terminus (amino acids 415 to 524) revealed three nonoverlapping antigenic sites in regions containing the residues (440)VPQVRKETRASSR(452) (site 1), (479)PEADTDPL(486) (site 2), and (520)DKDLL(524) (site 3). Among these, antigenic site 2 showed the strongest reactivity with hyperimmune anti-RPV bovine sera in a peptide enzyme-linked immunosorbent assay but did not react with hyperimmune caprine sera raised against peste-des-petits-ruminants virus, which is antigenically closely related to RPV. Identification of an immunodominant linear antigenic site at the carboxy terminus of the N protein may provide an antigen basis for designing diagnostics specific for RPV.
Choi
Kang-Seuk
KS
Foreign Animal Disease Research Division, National Veterinary Research and Quarantine Service, 480 Anyang-6-dong, Anyang, Kyonggi 430-824, Republic of Korea. choiks@nvrqs.go.kr
Nah
Jin-Ju
JJ
Ko
Young-Joon
YJ
Kang
Shien-Young
SY
Yoon
Kyoung-Jin
KJ
Joo
Yi-Seok
YS
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D001402:B-Lymphocytes / Q000276:immunology*; D015153:Blotting, Western; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D018604:Epitope Mapping; D005982:Glutathione Transferase; D006023:Glycoproteins / Q000276:immunology; D016056:Immunodominant Epitopes / Q000276:immunology*; D008565:Membrane Proteins; D019590:Nucleocapsid Proteins / Q000276:immunology*; D012302:Rinderpest virus / Q000276:immunology*; D014760:Viral Fusion Proteins / Q000276:immunology
D000914:Antibodies, Viral; C079748:F protein, Rinderpest virus; D006023:Glycoproteins; D016056:Immunodominant Epitopes; D008565:Membrane Proteins; D019590:Nucleocapsid Proteins; D014760:Viral Fusion Proteins; C481383:nucleocapsid protein, Rinderpest virus; D005982:Glutathione Transferase