Modulation of Mycobacterium tuberculosis DnaA protein-adenine-nucleotide interactions by acidic phospholipids.
The biochemical aspects of the initiation of DNA replication in Mycobacterium tuberculosis are unknown. To understand this process, we overproduced, purified and characterized the recombinant M. tuberculosis DnaA protein. The M. tuberculosis DnaA protein binds the origin of replication (oriC), ATP and ADP, and exhibited weak ATPase activity. ADP, after hydrolysis of ATP, remained strongly associated with DnaA and the exchange of ATP for bound ADP was weak. Vesicles prepared from acidic phospholipids, such as phosphatidylinositol, cardiolipin and phosphatidylglycerol, promoted dissociation of both ADP and ATP, whereas the neutral phospholipid phosphatidylethanolamine did not. The phospholipid-mediated dissociation of ATP was decreased in the presence of the M. tuberculosis oriC, whereas dissociation of ADP was stimulated in the presence of oriC. Acidic phospholipids in micelles, however, were not efficient in dissociating bound nucleotides from DnaA. Together, these results suggest that both polar head groups and membrane bilayer structure play an important role in M. tuberculosis DnaA-adenine-nucleotide interactions. We suggest that initiation of M. tuberculosis oriC involves intimate interactions between DnaA, adenine nucleotides and membrane phospholipids, and the latter helps to ensure that only the ATP form of the DnaA protein interacts continuously with oriC.
Yamamoto
Kohji
K
Biomedical Research, The University of Texas Health Center at Tyler, 11937 U.S. Hwy @ 271, Tyler, TX 75708-3154, U.S.A.
Muniruzzaman
Syed
S
Rajagopalan
Malini
M
Madiraju
Murty V V S
MV
D000227:Adenine Nucleotides / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008823:Micelles; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism*; D051738:Origin Recognition Complex; D010743:Phospholipids / Q000737:chemistry / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins
D000227:Adenine Nucleotides; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D008823:Micelles; C114650:OriC chromosomal replication origin; D051738:Origin Recognition Complex; D010743:Phospholipids; D011994:Recombinant Proteins; D014764:Viral Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate