Bacteriophage phi 29 DNA replication in vitro: participation of the terminal protein and the gene 2 product in elongation.

From phi 29-infected Bacillus subtilis cells, we have isolated a protein fraction which promotes in vitro replication of phi 29 DNA. This fraction catalyses both initiation and elongation, indicating that it contains the product of gene 3 (tp: terminal protein) and the product of gene 2 (gp2: probably a DNA polymerase), since initiation requires the two products (Blanco et al. 1983; Matsumoto et al. 1983). The fractions isolated from cells infected with temperature-sensitive (ts) mutants of gene 2 and gene 3 were thermolabile in both the initiation and elongation assays. When the pre-initiated material from the ts fractions of each mutant was heat-inactivated and mixed no complementation, restoring the elongation activity, was found. These results indicate: (i) tp and gp2 participate not only in the initiation but also in the elongation of phi 29 DNA replication, (ii) they probably function in tight physical association with each other.