Mapping of two antigenic domains on the NS3 protein of the pestivirus bovine viral diarrhea virus.
The immunodominant NS3 (p80) protein of the pestivirus bovine viral diarrhea virus (BVDV) functions as a serine protease and a RNA helicase. To identify antigenic domains of the BVDV NS3, a panel of monoclonal antibodies (mAbs) was tested against fragments of the protein expressed in E. coli. Two large overlapping NS3 fragments, A (amino acids [aa] 1-434) and B (aa 368-683) which together contain all NS3 sequences, were used to screen mAbs for reactivity. Two mAbs, 21.5.8 and 1.11.3, were reactive to fragment A (in ELISA only) and one mAb, 20.10.6, was reactive to fragment B (in ELISA and Western blotting). Further mapping demonstrated that the smallest fragment mAbs 21.5.8 and 1.11.3 bound to was comprised of aa 205-369 (domain A). In Western blotting, the smallest fragment reactive with mAb 20.10.6 was comprised of aa 368-549 (domain B). However, in indirect ELISA, mAb 20.10.6 also demonstrated high reactivity to a smaller fragment comprising aa 368-512 (domain B'). This indicated that the epitope of mAb 20.10.6 was conformational and not linear. Blocking ELISAs using these mAbs and type 1 and type 2 BVDV antisera demonstrated that an immunodominant region of the NS3 protein in cattle is defined by aa 205-549.
Deregt
Dirk
D
Lethbridge Laboratory (Animal Diseases Research Institute), Canadian Food Inspection Agency, P.O. Box 640, Lethbridge, Alta., Canada T1J 3Z4. deregtd@inspection.gc.ca
Dubovi
Edward J
EJ
Jolley
Michael E
ME
Nguyen
Phuong
P
Burton
Kimberley M
KM
Gilbert
Scott A
SA
D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral; D002417:Cattle; D001908:Diarrhea Viruses, Bovine Viral / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D016056:Immunodominant Epitopes / Q000032:analysis / Q000276:immunology*; D051379:Mice; D010447:Peptide Hydrolases / Q000276:immunology*; D020365:RNA Helicases / Q000276:immunology*; D017361:Viral Nonstructural Proteins / Q000276:immunology*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D016056:Immunodominant Epitopes; D010447:Peptide Hydrolases; D020365:RNA Helicases; D017361:Viral Nonstructural Proteins; C086730:p80 protein, bovine viral diarrhea virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases