Bidirectional replication from an internal ori site of the linear N15 plasmid DNA. Nucleic Acids Res

The prophage of coliphage N15 is not integrated into the chromosome but exists as a linear plasmid molecule with covalently closed hairpin ends (telo-meres). Upon infection the injected phage DNA circularizes via its cohesive ends. Then, a phage-encoded enzyme, protelomerase, cuts the circle and forms the hairpin telomeres. N15 protelomerase acts as a telomere-resolving enzyme during pro-phage DNA replication. We characterized the N15 replicon and found that replication of circular N15 miniplasmids requires only the repA gene, which encodes a multidomain protein homologous to rep-lication proteins of bacterial plasmids replicated by a theta-mechanism. Replication of a linear N15 mini-plasmid also requires the protelomerase gene and telomere regions. N15 prophage replication is initi-ated at an internal ori site located within repA and proceeds bidirectionally. Electron microscopy data suggest that after duplication of the left telomere, protelomerase cuts this site generating Y-shaped molecules. Full replication of the molecule and sub-sequent resolution of the right telomere then results in two linear plasmid molecules. N15 prophage replication thus appears to follow a mechanism that is distinct from that employed by eukaryotic replicons with this type of telomere and suggests the possibility of evolutionarily independent appear-ances of prokaryotic and eukaryotic replicons with covalently closed telomeres