No abstract

Reovirus guanylyltransferase, studied as a covalent enzyme-GMP intermediate, was used to guanylate appropriate acceptor molecules in vitro to produce authentic cap structures. Guanylyltransferase activity was associated with A2, the 140-kilodalton product of the L2 gene segment of reovirus serotypes 1 and 3. The proteins of reovirus are arranged as a double capsid structure. The inner capsid, or core, is composed of six viral proteins and contains the 10 double-stranded RNA (dsRNA) genome segments. All of the enzymes required for transcription and capping of viral mRNAs are associated with the viral core particle (1, 2, 7, 9, 11, 20, 22). Assignment of specific enzymatic activities to viral proteins has proven difficult, as solubilization of the core proteins invariably results in the loss of their enzymatic activities (31). Drayna and Fields (6) tentatively assigned the viral transcriptase activity to the core polypeptide A3 by using a genetic approach. In the present report, we used a direct biochemical approach to assign the reovirus guanylyltransferase activity to the core polypeptide X2. Shuman and Hurwitz (24) demonstrated that the guanylyltransferase reaction of vaccinia virus proceeds via a covalent enzyme-guanylate intermediate. A similar reaction mechanism has been demonstrated for the guanylyltransferase enzymes from a variety of organisms (10, 12, 15, 17, 19. 23, 28, 29, 32). Results obtained in our laboratory (35) and results obtained by Shatkin et al. (21) suggested that the * Corresponding author. This work was supported by grant MT-4594 from the Medical Research Council. Ottawa, Ontario. Canada. D.R.C. was a recipient of studentships from Fonds Formation de Chercheurs et Actions Concertees, Quebec. and the McGill McConnell Foundation. H.Z. was a recipient of a studentship from Fonds de la Recherche en Sante du Quebec.