The DnaA N-terminal domain interacts with Hda to facilitate replicase clamp-mediated inactivation of DnaA

Masayuki Su'etsugu, Yuji Harada, Kenji Keyamura, Chika Matsunaga, Kazutoshi Kasho, Yoshito Abe, Tadashi Ueda, Tsutomu Katayama

Research output: Contribution to journalArticlepeer-review

14 Citations (Scopus)


DnaA activity for replication initiation of the Escherichia coli chromosome is negatively regulated by feedback from the DNA-loaded form of the replicase clamp. In this process, called RIDA (regulatory inactivation of DnaA), ATP-bound DnaA transiently assembles into a complex consisting of Hda and the DNA-clamp, which promotes inter-AAA+ domain association between Hda and DnaA and stimulates hydrolysis of DnaA-bound ATP, producing inactive ADP-DnaA. Using a truncated DnaA mutant, we previously demonstrated that the DnaA N-terminal domain is involved in RIDA. However, the precise role of the N-terminal domain in RIDA has remained largely unclear. Here, we used an in vitro reconstituted system to demonstrate that the Asn-44 residue in the N-terminal domain of DnaA is crucial for RIDA but not for replication initiation. Moreover, an assay termed PDAX (pull-down after cross-linking) revealed an unstable interaction between a DnaA-N44A mutant and Hda. In vivo, this mutant exhibited an increase in the cellular level of ATP-bound DnaA. These results establish a model in which interaction between DnaA Asn-44 and Hda stabilizes the association between the AAA+ domains of DnaA and Hda to facilitate DnaA-ATP hydrolysis during RIDA.

Original languageEnglish
Pages (from-to)3183-3195
Number of pages13
JournalEnvironmental Microbiology
Issue number12
Publication statusPublished - Dec 2013

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Ecology, Evolution, Behavior and Systematics


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