The N-terminal half of the brome mosaic virus 1a protein has RNA capping-associated activities: Specificity for GTP and S-adenosylmethionine

The N-terminal half of the brome mosaic virus (BMV) 1a replication-associat ed protein contains sequence motifs found in RNA methyltransferases. We dem onstrate that recombinant BMV methyltransferase-like (MT) domain expressed In Escherichia coil forms an adduct with a guanine nucleotide in a reaction that requires S-adenosylmethionine (AdoMet) and divalent cations. Moieties in GTP and AdoMet required for adduct formation were determined using a co mpetition assay and chemical analogues. In the guanine nucleotide the ribos e 2' hydroxyl, the triphosphates, the base C6 keto group, and possibly the N1 imine are required. In AdoMet, the methyl group and the ability to trans fer a methyl group to guanine nucleotide were demonstrated to be required f or adduct formation. The effects of methyltransferase inhibitors on viral R NA synthesis was determined using an in vitro RNA synthesis assay. These re sults are consistent with the previously reported activities of alphaviral nsP1 methyltransferase protein and identify the chemical moieties required for the BMV methyltransferase activity. (C) 1999 Academic Press.