Nucleotide sequence and expression in Escherichia coli of the gene encoding the nonstructural protein NCVP2 of bovine rotavirus.

Cloned DNA copy of rotavirus genome segment 5 from bovine rotavirus RF strain has been used to determine the nucleotide sequence of the gene that encodes for the nonstructural viral protein NCVP2. The sequence data indicated that segment 5 consists of 1581 base pairs and is A + T rich (66%). The positive strand of segment 5 contains a single open reading frame that extends 491 codons and possesses 5'- and 3'-terminal untranslated regions of 32 and 73 base pairs, respectively. The first AUG conforms to the Kozak consensus sequence and if utilized, would yield a protein having a calculated molecular weight of 58,654, slightly higher than the apparent molecular weight of NCVP2 (MW 54,000). Although it is not evident whether the gene product is glycosylated, four potential glycosylation sites were found at positions 50, 168, 403, and 438. NCVP2 has been expressed in Escherichia coli using the inducible expression vector pKK233-2. Following IPTG induction high levels of full-length nonfused proteins were synthesized and accumulated in induced cells.