Isolation and enzymatic characterization of protein λ2, the reovirus guanylyltransferase

Abstract

Protein λ2 of reovirus serotype 3 has been purified to homogeneity from extracts of cells infected with hybrid vaccinia virus strain WR into whose TK gene of the reovirus L2 genome segment under the control of the CPV ATI protein gene promoter had been inserted. Protein λ2 is formed in large amounts (final purification factor about 180) as a monomer that shows no tendency to pentamerize into the reovirus core projections/spikes. Isolated protein λ2 is reversibly guanylylated by GTP (that is, it carries out the GTP- PP i exchange reaction) and can transfer the -GMP moiety to GTP to form GppppG, to GDP to form GpppG, and to 5′-pp-terminated RNA to form GpppG- caps. These studies confirm previous studies on reovirus cores that indicated that protein λ2 is the reovirus guanylyltransferase. Protein λ2 possesses neither nucleoside nor RNA triphosphatase activities, nor methyltransferase activities; thus it is the reovirus capping enzyme, but provides neither the required 5′-ppG-terminated substrate nor does it methylate the cap structure. These must be functions of λ2 pentamers or of other individual or complexed components of reovirus cores.