Rifampicin-resistant initiation of DNA synthesis on the isolated strands of ColE plasmid DNA.
The opposite strands of the ColE1 and ColE3 plasmids were isolated as circular single-stranded DNA molecules. These molecules were compared with M13 and phi X174 viral DNA with respect to their capacity to function as templates for in vitro DNA synthesis by a replication enzyme fraction from Escherichia coli. It was found for both ColE plasmids that the conversion of H as well as L strands to duplex DNA molecules closely resembles phi X174 complementary strand synthesis and occurs by a rifampicin-resistant priming mechanism involving the dnaB, dnaC, and dnaG gene products. Restriction analysis of partially double-stranded intermediates indicates that preferred start sites for DNA synthesis are present on both strands of the ColE1 HaeII-C fragment. Inspection of the nucleotide sequence of this region reveals structural similarities with the origin of phi X174 complementary strand synthesis. We propose that the rifampicin-resistant initiation site (rri) in the ColE1 L strand is required for the priming of discontinuous lagging strand synthesis during vegetative replication and that the rri site in the H strand is involved in the initiation of L strand synthesis during conjugative transfer.
D004261:DNA Replication / Q000187:drug effects*; D004262:DNA Restriction Enzymes; D004270:DNA, Circular / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D009492:Neurospora crassa / Q000235:genetics; D010957:Plasmids; D012293:Rifampin / Q000494:pharmacology*
D004270:DNA, Circular; D004277:DNA, Single-Stranded; D004262:DNA Restriction Enzymes; D012293:Rifampin