A novel mode of DnaA-DnaA interaction promotes ADP dissociation for reactivation of replication initiation activity

Nucleic Acids Res. 2019 Dec 2;47(21):11209-11224. doi: 10.1093/nar/gkz795.


ATP-DnaA is temporally increased to initiate replication during the cell cycle. Two chromosomal loci, DARS (DnaA-reactivating sequences) 1 and 2, promote ATP-DnaA production by nucleotide exchange of ADP-DnaA for timely initiation. ADP-DnaA complexes are constructed on DARS1 and DARS2, bearing a cluster of three DnaA-binding sequences (DnaA boxes I-III), promoting ADP dissociation. Although DnaA has an AAA+ domain, which ordinarily directs construction of oligomers in a head-to-tail manner, DnaA boxes I and II are oriented oppositely. In this study, we constructed a structural model of a head-to-head dimer of DnaA AAA+ domains, and analyzed residues residing on the interface of the model dimer. Gln208 was specifically required for DARS-dependent ADP dissociation in vitro, and in vivo analysis yielded consistent results. Additionally, ADP release from DnaA protomers bound to DnaA boxes I and II was dependent on Gln208 of the DnaA protomers, and DnaA box III-bound DnaA did not release ADP nor require Gln208 for ADP dissociation by DARS-DnaA complexes. Based on these and other findings, we propose a model for DARS-DnaA complex dynamics during ADP dissociation, and provide novel insight into the regulatory mechanisms of DnaA and the interaction modes of AAA+ domains.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Diphosphate / metabolism*
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • DNA Replication / physiology*
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / metabolism
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Models, Molecular
  • Mutant Proteins / chemistry
  • Mutant Proteins / metabolism
  • Origin Recognition Complex / chemistry
  • Origin Recognition Complex / metabolism*
  • Protein Binding
  • Protein Multimerization / physiology*
  • Protein Structure, Quaternary


  • Bacterial Proteins
  • DNA, Bacterial
  • DNA-Binding Proteins
  • DnaA protein, Bacteria
  • Mutant Proteins
  • OriC chromosomal replication origin
  • Origin Recognition Complex
  • Adenosine Diphosphate