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The intrinsic ATPase activity of Mycobacterium tuberculosis DnaA promotes rapid oligomerization of DnaA on oriC
Author(s) -
Madiraju Murty V. V. S.,
Moomey Meredith,
Neuenschwander Pierre F.,
Muniruzzaman Syed,
Yamamoto Kohji,
Grimwade Julia E.,
Rajagopalan Malini
Publication year - 2006
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2006.05068.x
Subject(s) - dnaa , dnab helicase , biology , origin of replication , dna replication , atpase , dna , mutant , escherichia coli , biochemistry , microbiology and biotechnology , biophysics , gene , enzyme , rna , helicase
Summary 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 DnaA TB promoted rapid oligomerization on oriC. In support, we found that a recombinant mutant DnaA TB 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 DnaA TB bound to DnaA boxes similarly with ATP or ADP. DnaA TB 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.