Initiator protein DnaA of Escherichia coli is a negative replication regulator of linear phage-plasmid N15

Abstract

Temperate bacteriophage N 15 in the lysogenic state is incapable of integrating in the chromosome of Escherichia coli and represents a linear plasmid with covalently closed ends. The phage repA gene, the product of which possesses activities of primase and helicase, ensures replication of N15 DNA. The ori site of initiation of N15 replication contains binding sites for RepA and a potential site of binding the bacterial initiator protein DnaA. It was shown in our work that replication of miniplasmids based on N15 replicon as well as replication of N15 DNA during lytic growth do not depend on DnaA. Moreover, introducing mutations into the potential DnaA binding site increases the copy number of circular and linear miniplasmids that contain repA gene. These data suggest that DnaA is a negative rather than positive regulator of phage N15 replication. This is assumed to be caused by properties of interaction between RepA and DnaA during initiation of N15 replication or by transcriptional silencing of repA gene due to the binding of DnaA to the ori site located within the coding repA sequence.