title: Cooperative binding mode of the inhibitors of R6K replication, pi dimers.
abstract: The replication initiator protein, pi, plays an essential role in the initiation of plasmid R6K replication. Both monomers and dimers of pi bind to iterons in the gamma origin of plasmid R6K, yet monomers facilitate open complex formation, while dimers, the predominant form in the cell, do not. Consequently, pi monomers activate replication, while pi dimers inhibit replication. Recently, it was shown that the monomeric form of pi binds multiple tandem iterons in a strongly cooperative fashion, which might explain how monomers outcompete dimers for replication initiation when plasmid copy number and pi supply are low. Here, we examine cooperative binding of pi dimers and explore the role that these interactions may have in the inactivation of gamma origin. To examine pi dimer/iteron interactions in the absence of competing pi monomer/iteron interactions using wild-type pi, constructs were made with key base changes to each iteron that eliminate pi monomer binding yet have no impact on pi dimer binding. Our results indicate that, in the absence of pi monomers, pi dimers bind with greater cooperativity to alternate iterons than to adjacent iterons, thus preferentially leaving intervening iterons unbound and the origin unsaturated. We discuss new insights into plasmid replication control by pi dimers.
authors.affiliation: Department of Bacteriology, University of Wisconsin-Madison, 1550 Linden Drive, Madison, WI 53706, USA.
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Bowers
authors.forename: Marcin
authors.initials: M
authors.lastname: Filutowicz
mesh_terms: D004265:DNA Helicases / Q000235:genetics*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics*; D019281:Dimerization; D018628:Gene Dosage; D009154:Mutation; D009690:Nucleic Acid Conformation; D010957:Plasmids; D018741:Replication Origin; D015534:Trans-Activators / Q000235:genetics*
chemical_list: D004268:DNA-Binding Proteins; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases