title: Mapping of epitopes and structural analysis of antigenic sites in the nucleoprotein of rabies virus.
abstract: Linear epitopes on the rabies virus nucleoprotein (N) recognized by six MAbs raised against antigenic sites I (MAbs 6-4, 12-2 and 13-27) and IV (MAbs 6-9, 7-12 and 8-1) were investigated. Based on our previous studies on sites I and IV, 24 consecutively overlapping octapeptides and N- and C-terminal-deleted mutant N proteins were prepared. Results showed that all three site I epitopes studied and two site IV epitopes (for MAbs 8-1 and 6-9) mapped to aa 358-367, and that the other site IV epitope of MAb 7-12 mapped to aa 375-383. Tests using chimeric and truncated proteins showed that MAb 8-1 also requires the N-terminal sequence of the N protein to recognize its binding region more efficiently. Immunofluorescence studies demonstrated that all three site I-specific MAbs and one site IV-specific MAb (7-12) stained the N antigen that was diffusely distributed in the whole cytoplasm; the other two site IV-specific MAbs (6-9 and 8-1) detected only the N antigen in the cytoplasmic inclusion bodies (CIB). An antigenic site II-specific MAb (6-17) also detected CIB-associated N antigen alone. Furthermore, the level of diffuse N antigens decreased after treatment of infected cells with cycloheximide. These results suggest that epitopes at site I are expressed on the immature form of the N protein, but epitope structures of site IV MAbs 6-9 and 8-1 are created and/or exposed only after maturation of the N protein.
authors.affiliation: Department of Veterinary Public Health, Faculty of Agriculture, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Goto
authors.forename: N
authors.initials: N
authors.lastname: Minamoto
authors.forename: H
authors.initials: H
authors.lastname: Ito
authors.forename: N
authors.initials: N
authors.lastname: Ito
authors.forename: M
authors.initials: M
authors.lastname: Sugiyama
authors.forename: T
authors.initials: T
authors.lastname: Kinjo
authors.forename: A
authors.initials: A
authors.lastname: Kawai
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015153:Blotting, Western; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D019084:Fluorescent Antibody Technique, Indirect; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D009698:Nucleoproteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D010957:Plasmids / Q000235:genetics; D011820:Rabies virus / Q000235:genetics / Q000276:immunology* / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism; D017384:Sequence Deletion
chemical_list: D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D010455:Peptides; D011993:Recombinant Fusion Proteins