In vitro replication of bacteriophage GA RNA. Subunit structure and catalytic properties of GA replicase

An RNA replicase of GA phage, one of the Group II RNA phages, was isolated and purified to a homogeneous state. By SDS polyacrylamide gel analysis, the purified GA replicase was found to contain four different subunits, numbered I, II, III, and IV, the molecular weights of which were 74,000, 60,000, 47,000, and 36,000, respectively. Three of them, I, III, and IV, proved to be host-coded proteins, ribosomal protein S1 (I), and elongation factors Tu (III) and Ts (IV) of protein biosynthesis, respectively. On a phosphocellulose column, the RNA replicase was separated into two components: One composed of subunits I and II, and the other composed of subunits III and IV. Each component alone had no replicase activity. However, when the two components were combined at 0 degree C, 60% of the replicase activity was restored within 10 min. The purified GA replicase catalyzed the GA phage RNA-directed synthesis of template-size RNA. However, the maximum level of product RNA synthesized was less than 20% of the amount of template RNA added. RNA-RNA hybridization experiments indicated that the product RNA included only the RNA strand complementary to the template RNA, and not the viral strand.