Mutations in a herpes simplex virus type 1 origin that inhibit interaction with origin-binding protein also inhibit DNA replication

The herpes simplex virus type 1 genome contains three origins of replication: Ori_L and a diploid Ori_S. The origin-binding protein, the product of the UL9 gene, interacts with two sites within Ori_S, box I and box II. A third site, box III, which is homologous to boxes I and II, may also be a binding site for the origin-binding protein. Mutations in these three sites significantly reduce Ori_S-directed plasmid replication measured in transient replication assays. The reduction in replication efficiency of the mutants correlates well with the decrease in the ability to bind to the origin-binding protein, as determined by Elias et al. (P. Elias, C. M. Gustafsson, and O. Hammarsten, J. Biol. Chem. 265:17167-17173, 1990). The effect of multiple mutations in boxes I, II, and HI on plasmid replication suggests that there are multiple binding sites in Ori_S for the origin-binding protein. These studies indicate that proper interaction of the origin-binding protein with the Ori_S sequence is essential for Ori_S-directed DNA replication.