The intrinsic ATPase activity of Mycobacterium tuberculosis DnaA promotes rapid oligomerization of DnaA on oriC

Murty V.V.S. Madiraju, Meredith Moomey, Pierre F. Neuenschwander, Syed Muniruzzaman, Kohji Yamamoto, Julia E. Grimwade, Malini Rajagopalan

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Abstract

Oligomerization of the initiator protein, DnaA, on the origin of replication (oriC) is crucial for initiation of DNA replication. Studies in Escherichia coli (Gram-negative) have revealed that binding of DnaA to ATP, but not hydrolysis of ATP, is sufficient to promote DnaA binding, oligomerization and DNA strand separation. To begin understanding the initial events involved in the initiation of DNA replication in Mycobacterium tuberculosis (Gram-positive), we investigated interactions of M. tuberculosis DnaA (DnaA TB) with oriC using surface plasmon resonance in the presence of ATP and ADP. We provide evidence that, in contrast to what is observed in E. coli, ATPase activity of DnaATB promoted rapid oligomerization on oriC. In support, we found that a recombinant mutant DnaATB proficient in binding to ATP, but deficient in ATPase activity, did not oligomerize as rapidly. The corresponding mutation in the dnaA gene of M. tuberculosis resulted in non-viability, presumably due to a defect in oriC-DnaA interactions. Dimethy sulphate (DMS) footprinting experiments revealed that DnaATB bound to DnaA boxes similarly with ATP or ADP. DnaATB binding to individual DnaA boxes revealed that rapid oligomerization on oriC is triggered only after the initial interaction of DnaA with individual DnaA boxes. We propose that ATPase activity enables the DnaA protomers on oriC to rapidly form oligomeric complexes competent for replication initiation.

Original languageEnglish
Pages (from-to)1876-1890
Number of pages15
JournalMolecular Microbiology
Volume59
Issue number6
DOIs
Publication statusPublished - Mar 2006

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

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    Madiraju, M. V. V. S., Moomey, M., Neuenschwander, P. F., Muniruzzaman, S., Yamamoto, K., Grimwade, J. E., & Rajagopalan, M. (2006). The intrinsic ATPase activity of Mycobacterium tuberculosis DnaA promotes rapid oligomerization of DnaA on oriC. Molecular Microbiology, 59(6), 1876-1890. https://doi.org/10.1111/j.1365-2958.2006.05068.x