Regulatory dynamics in the ternary DnaA complex for initiation of chromosomal replication in Escherichia coli

Yukari Sakiyama, Kazutoshi Kasho, Yasunori Noguchi, Hironori Kawakami, Tsutomu Katayama

Research output: Contribution to journalArticlepeer-review

16 Citations (Scopus)

Abstract

In Escherichia coli, the level of the ATP-DnaA initiator is increased temporarily at the time of replication initiation. The replication origin, oriC, contains a duplex-unwinding element (DUE) flanking a DnaAoligomerization region (DOR), which includes twelve DnaA-binding sites (DnaA boxes) and the DNAbending protein IHF-binding site (IBS). Although complexes of IHF and ATP-DnaA assembly on the DOR unwind the DUE, the configuration of the crucial nucleoprotein complexes remains elusive. To resolve this, we analyzed individual DnaA protomers in the complex and here demonstrate that the DUE- DnaA-box-R1-IBS-DnaA-box-R5M region is essential for DUE unwinding. R5M-bound ATP-DnaA predominantly promotes ATP-DnaA assembly on the DUE-proximal DOR, and R1-bound DnaA has a supporting role. This mechanism might support timely assembly of ATP-DnaA on oriC. DnaA protomers bound to R1 and R5M directly bind to the unwound DUE strand, which is crucial in replication initiation. Data from in vivo experiments support these results. We propose that the DnaA assembly on the IHF-bent DOR directly binds to the unwound DUE strand, and timely formation of this ternary complex regulates replication initiation. Structural features of oriC support the idea that these mechanisms for DUE unwinding are fundamentally conserved in various bacterial species including pathogens.

Original languageEnglish
Pages (from-to)12354-12373
Number of pages20
JournalNucleic acids research
Volume45
Issue number21
DOIs
Publication statusPublished - Dec 1 2017

All Science Journal Classification (ASJC) codes

  • Genetics

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