Identification of the amino acid residue involved in rabbit hemorrhagic disease virus VPg uridylylation

The virus genome-linked protein (VPg) coding region from rabbit hemorrhagic disease virus (RHDV) (isolate AST/89) was expressed in Escherichia coli by using a glutathione S-transferase-based vector. The recombinant polypeptid e could be purified in good yields and was uridylylated in vitro from [alph a-P-32]UTP in a reaction catalyzed by the recombinant RNA-dependent RNA pol ymerase from RHDV in the absence of added template RNA. The use of deletion and point mutants allowed the identification of Tyr-21 as the residue invo lved in uridylylation and consequently in the linkage between VPg and the v iral genome. These data constitute the first report on the identity of the amino acid residue involved in VPg uridylylation in a member of the Caliciv iridae family.