Sequence-independent DNA binding activity of DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli.
The DnaA protein specifically binds to the origin of chromosomal DNA replication and initiates DNA synthesis. In addition to this sequence-specific DNA binding, DnaA protein binds to DNA in a sequence-independent manner. We here compared the two DNA binding activities. Binding of ATP and ADP to DnaA inhibited the sequence-independent DNA binding, but not sequence-specific binding. Sequence-independent DNA binding, but not sequence-specific binding, required incubation at high temperatures. Mutations in the C-terminal domain affected the sequence-independent DNA binding activity less drastically than they did the sequence-specific binding. On the other hand, the mutant DnaA433, which has mutations in a membrane-binding domain (K327 to I344) was inert for sequence-independent binding, but could bind specifically to DNA. These results suggest that the two DNA binding activities involve different domains and perform different functions from each other in Escherichia coli cells.
Makise
Masaki
M
Faculty of Pharmaceutical Sciences, Okayama University, and PRESTO, Japan Science and Technology Corporation, Okayama 700-8530, Japan.
Tsuchiya
Tomofusa
T
Mizushima
Tohru
T
D000227:Adenine Nucleotides / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000187:drug effects; D002876:Chromosomes, Bacterial; D004252:DNA Mutational Analysis; D004261:DNA Replication / Q000235:genetics* / Q000502:physiology; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D013696:Temperature
D000227:Adenine Nucleotides; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D000255:Adenosine Triphosphate