Topological characterization of the DnaA-oriC complex using single-molecule nanomanipuation

In most bacteria, the timing and synchrony of initiation of chromosomal replication are determined by the binding of the AAA(+) protein DnaA to a set of high- and low-affinity sites found within the origin of chromosomal replication (oriC). Despite the large amount of information on the role and reg...

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Bibliographic Details
Published in:Nucleic acids research 2012-08, Vol.40 (15), p.7375-7383
Main Authors: Zorman, Sylvain, Seitz, H, Sclavi, B, Strick, T R
Format: Article
Language:English
Subjects:
Adenosine Triphosphate
Adenosine Triphosphate - metabolism
Bacterial Proteins
Bacterial Proteins - chemistry
Bacterial Proteins - metabolism
Biochemistry, Molecular Biology
Biomechanical Phenomena
Biomechanics
Crystal structure
DNA
DNA, Bacterial
DNA, Bacterial - chemistry
DNA-Binding Proteins
DNA-Binding Proteins - chemistry
DNA-Binding Proteins - metabolism
Escherichia coli
Escherichia coli - genetics
Genome Integrity, Repair and
Handedness
Life Sciences
Mutation
Origin Recognition Complex
Origin Recognition Complex - chemistry
Origin Recognition Complex - genetics
Replication
Replication initiation
Replication Origin
Replication origins
Unwinding
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Summary:In most bacteria, the timing and synchrony of initiation of chromosomal replication are determined by the binding of the AAA(+) protein DnaA to a set of high- and low-affinity sites found within the origin of chromosomal replication (oriC). Despite the large amount of information on the role and regulation of DnaA, the actual structure of the DnaA-oriC complex and the mechanism by which it primes the origin for the initiation of replication remain unclear. In this study, we have performed magnetic tweezers experiments to investigate the structural properties of the DnaA-oriC complex. We show that the DnaA-ATP-oriC complex adopts a right-handed helical conformation involving a variable amount of DNA and protein whose features fit qualitatively as well as quantitatively with an existing model based on the crystal structure of a truncated DnaA tetramer obtained in the absence of DNA. We also investigate the topological effect of oriC's DNA unwinding element.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/gks371