title: Nucleotide sequence of bovine rotavirus gene 1 and expression of the gene product in baculovirus.
abstract: The nucleotide sequence of the gene that encodes for the structural viral protein VP1 of bovine rotavirus (RF strain) has been determined. The sequence data indicate that segment 1 contains 3302 bp and is A + T rich (65.3%). The positive strand of segment 1 contains a single open reading frame that extends 1088 codons and possesses 5'- and 3'-terminal untranslated regions of 18 and 20 bp, respectively. The first AUG conforms to the Kozak consensus sequence and if utilized, would yield a protein having a calculated molecular weight of 124,847, very close to the apparent molecular weight of VP1 (M.W. 125,000). The deduced amino acid sequence presents significant similarities with RNA-dependent RNA polymerase of several RNA viruses. VP1 was also synthesized in baculovirus using two transfer vecors: pAC461 and pVL941. Following infection of Sf9 cells with a recombinant baculovirus, a full-length nonfusion protein was synthesised which shares properties with authentic VP1 made in monkey kidney cells. The level of VP1 synthesis was about 10-fold higher when the baculovirus recombinant was derived from the pVL941 transfer vector. In that case, VP1 was expressed in yields approximately equivalent to 10% of the cellular protein. The recombinant protein was immunoprecipitated by hyperimmune serum raised against purified rotavirus. It also was immunogenic; a hyperimmune serum made in guinea pigs reacted with VP1 using immunoprecipitation and Western blot. This serum did not possess neutralization activity.
authors.affiliation: Station de Virologie et d'Immunologie Moléculaires, INRA, C.R.J. Domaine de Vilvert, Jouy-en-Josas, France.
authors.forename: J
authors.initials: J
authors.lastname: Cohen
authors.forename: A
authors.initials: A
authors.lastname: Charpilienne
authors.forename: S
authors.initials: S
authors.lastname: Chilmonczyk
authors.forename: M K
authors.initials: MK
authors.lastname: Estes
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D005786:Gene Expression Regulation; D005814:Genes, Viral; D007304:Insect Viruses / Q000235:genetics; D008969:Molecular Sequence Data; D012401:Rotavirus / Q000201:enzymology / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology; D015678:Viral Structural Proteins
chemical_list: D014764:Viral Proteins; D015678:Viral Structural Proteins; D012321:DNA-Directed RNA Polymerases