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Monomer/dimer ratios of replication protein modulate the DNA strand-opening in a replication origin
- Source :
- Journal of molecular biology. 306(5)
- Publication Year :
- 2001
-
Abstract
- DNA opening is an essential step in the initiation of replication via the Cairns mode of replication. The opening reaction was investigated in a gamma ori system by using hyperactive variants of plasmid R6K-encoded initiator protein, pi. Reactivity to KMnO4 (indicative of opening) within gamma ori DNA occurred in both strands of a superhelical template upon the combined addition of wt pi, DnaA and integration host factor (IHF), each protein known to specifically bind gamma ori. IHF, examined singly, enhanced reactivity to KMnO4. The IHF-dependent reactive residues, however, are distinct from those dependent on pi (wt and hyperactive variants). Remarkably, the DNA helix opening does not require IHF and/or DnaA when hyperactive variants of pi were used instead of wt protein. We present three lines of evidence consistent with the hypothesis that DNA strand separation is facilitated by pi monomers despite the fact that both monomers and dimers of the protein can bind to iterons (pi binding sites). Taken together, our data suggest that pi elicits its ability to modulate plasmid copy number at the DNA helix-opening step.
- Subjects :
- DNA Replication
Gene Dosage
Biology
Polymerase Chain Reaction
chemistry.chemical_compound
Plasmid
Bacterial Proteins
Potassium Permanganate
Structural Biology
Escherichia coli
Reactivity (chemistry)
Binding site
Molecular Biology
Host factor
DNA Primers
Binding Sites
Base Sequence
Molecular biology
AT Rich Sequence
DnaA
DNA-Binding Proteins
Monomer
chemistry
Amino Acid Substitution
Helix
Biophysics
bacteria
Nucleic Acid Conformation
Replicon
DNA
Plasmids
Subjects
Details
- ISSN :
- 00222836
- Volume :
- 306
- Issue :
- 5
- Database :
- OpenAIRE
- Journal :
- Journal of molecular biology
- Accession number :
- edsair.doi.dedup.....fdec5ac46b11eb8abbe6bb99c7eb62d3