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Specialized DnaA binding sites in Escherichia coli oriC are targeted by multiple mechanisms regulating initiation of DNA replication
Author(s) -
Leonard Alan C,
Grimwade Julia E,
Nievera Christian,
Torgue Julien JC
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a511-a
Subject(s) - dnaa , dna replication , dna , microbiology and biotechnology , biology , dnab helicase , origin of replication , origin recognition complex , biophysics , biochemistry , helicase , eukaryotic dna replication , rna , gene
Regulatory mechanisms must exist in all cells to ensure new rounds of DNA synthesis are properly timed and limited to once per cell cycle. A critical step in initiation of DNA replication is stable unwinding of replication origin DNA. In E. coli , this is achieved by staged assembly of nucleoprotein complexes (orisomes) comprising AAA+ initiator DnaA and bending proteins Fis and IHF. In vitro and in vivo dimethyl sulfate footprinting studies have revealed specialized low affinity DnaA binding sites that must be filled before origin DNA can be unwound. DnaA binding to these sites is restricted to the time of initiation, regulated by Fis, IHF, and levels of DnaA‐ATP. In contrast, DnaA is bound to higher affinity sites throughout the cell cycle. After initiation, in E. coli three known inactivating mechanisms prevent reinitiation. We have found that these inactivating mechanisms reduce DnaA binding to the same specialized sites in oriC that determined orisome assembly, and so prevent reformation of the strand separated complex. We propose that the weakest DnaA‐ oriC interactions are targets for multiple mechanisms that promote or inhibit assembly of the unwound complex and ensure properly timed initiation during the cell cycle. NIH GM54042