Baculovirus expression of the ADRV gene 5 encoded protein produces an oligomerized, antigenic, and immunogenic VP6 protein.
Adult diarrheal rotavirus (ADRV) is a currently noncultivatable group B human rotavirus responsible for epidemic outbreaks of gastroenteritis in China. Gene segment 5 of ADRV encodes the major inner capsid protein, VP6. ADRV gene 5 was inserted into a recombinant baculovirus by homologous recombination between baculovirus shuttle plasmid pACYM1-AD5 and AcNPV genomic DNA. Baculovirus recombinants were selected visually and plaque purified and VP6 expression was detected by Coomassie staining of PAGE-separated proteins. The baculovirus-expressed gene 5 polypeptide is 44 kDa, the same as for the major inner capsid protein present on EDTA-treated ADRV virions and in vitro-expressed VP6 protein. The expressed protein is oligomeric and in the absence of reducing agents multimerizes to apparent trimer, hexamer, and greater molecular mass as assayed by SDS-PAGE. The VP6 protein is immunoprecipitable by hyperimmune serum to ADRV, human ADRV convalescent serum, by a group B-specific monoclonal antibody and by porcine group B rotavirus infection serum. The baculovirus-expressed protein is immunogenic and antibodies to the expressed protein recognize ADRV virions. The ADRV VP6 protein should be useful for developing diagnostic assays for serum antibodies to group B rotavirus as well as for generating hyperimmune serum and monoclonal antibodies for detecting viral antigen from ADRV and other group B rotaviruses.
Department of Veterans Affairs, Northport VA Medical Center, New York 11733.
E R
ER
Mackow
M E
ME
Fay
R
R
Werner-Eckert
T
T
Hung
Z J
ZJ
Wang
G
G
Chen
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D016367:Baculoviridae; D002213:Capsid / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D036022:Capsid Proteins; D015870:Gene Expression / Q000502:physiology; D005814:Genes, Viral / Q000502:physiology; D005822:Genetic Vectors; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology; D012401:Rotavirus / Q000737:chemistry*
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D036022:Capsid Proteins; D011994:Recombinant Proteins; C061332:VP6 protein, Rotavirus