DnaA protein

In E. coli cells, DNA replication starts at a specific site called oriC. The oriC locus contains only 245 base pairs. Similar sequences are responsible for initiating the synthesis of plasmid and bacteriophage DNA. The oriC nucleotide sequence binds several units of the tetrameric form of the dnaA protein. This protein is named for the gene that encodes it. The dnaB and dnaC proteins then bind to the complex. As a result of binding these proteins, a portion of the helical DNA is unwound. This forces the rest of the DNA into a left-handed double helix that wraps around the proteins to give a structure... 

Tandem array of Binding sites for DnaA protein,... 

FIGURE 25-12 Model for initiation of replication at the E. coli origin, oriC. (D About 20 DnaA protein molecules, each with a bound ATP, bind at the four 9 bp repeats. The DNA is wrapped around this complex. The three A=T-rich 13 bp repeats are denatured sequentially. (3) Hexamers of the DnaB protein bind to each strand, with the aid of DnaC protein. The DnaB helicase activity further unwinds the DNA in preparation for priming and DNA synthesis. 

DnaA protein 52,000 1 Recognizes ori sequence opens duplex at specific sites in... 

Immediately after replication, the DNA is hemi-methylated the parent strands have methylated oriC sequences but the newly synthesized strands do not. The hemimethylated oriC sequences are now sequestered for a period by interaction with the plasma membrane (the mechanism is unknown). After a time, oriC is released from the plasma membrane, and it must be fully methylated by Dam methylase before it can again bind DnaA. Regulation of initiation also involves the slow hydrolysis of ATP by DnaA protein, which cycles the pro-... 

A group of proteins form the prepriming complex. They recognize the origin of replication (dnaA protein), maintain the separation of the parental strands (single-stranded DNA-binding proteins), and unwind the double helix ahead of the advancing replication fork (helicase). 

Following the binding of the dnaA protein the hexameric helicase dnaB (Fig. 27-15) is loaded onto the adjacent DNA in the region of the 13-residue repeated... 

Figure 27-19 Hypothetical scheme for initiation of bidirectional replication in E. coli. The closed boxes R1 through R5 represent the 9-residue recognition sequences for the E. coli dnaA protein. The open boxes 1, 2, and 3 represent the three 13-residue repeats, possible sites for binding of the dnaB-dnaC protein complex. From McMacken et al.m Redrawn in simplified form.

It has long been postulated that the bacterial chromosome is attached to the plasma membrane. At least one such attachment site may be at or near the origin of replication.393 Furthermore, the exchange of ADP for ATP in the dnaA protein is catalyzed specifically by cardiolipin and phosphati-dylglycerol containing the unsaturated oleic acid.386 393 Inositol polyphosphates may also play a role 394... 

Initiation sequences contain a set of repeated sequences, which bind the essential initiator protein, DnaA. The DnaA protein opens the helix to make a short region of separated strands. Then a specialized single-strand binding protein binds to the DNA strands to keep them apart. This process makes a template, but replication can t happen because no primer yet exists. See Figure 8-10b. 

Presumably, such a mechanism calibrates the balance of ATP to ADP within the cell. During active replication, some 1,000-2,000 DnaA proteins are present in the cell, and occasionally as many as 10,000 (Donachie and Blakely 2003). These bind to either ATP or ADP, which in turn enables thermodynamic control over bacterial replication. This is because the Gibbs free energy of ATP hydrolysis depends on the ratio of ATP to ADP, not their total... 

Kitagawa R, Mitsuki H, Okazaki T, Ogawa T (1996) A novel DnaA protein-binding site at 94.7 min on the Escherichia coli chromosome. Mol Microbiol 19 1137-1147 Kleiber M (1961) The fire of life. Wiley, New York... 

Figure 27.25. Origin of Replication in E. coli. OriC has a length of 245 hp. It contains a tandem array of three nearly identical 13-nucleotide sequences (green) and four binding sites (yellow) for the dnaA protein. The relative orientations of the four dnaA sites are denoted hy arrows.

Location of A-T rich sequences and the four 9-base-pair repeats within OriC. (B). Model showing how DnaA protein binding to the 9-base-pair repeats leads to melting of the A-T rich sequence and formation of the two bidirectional replication forks. 

Binding of the yeast ORC proteins to the ORE DNA sequence is similar to the binding of E. coli DnaA proteins to the 9-base-pair repeat of OriC. A second similarity between yeast and E. coli is the presence of an A-T rich... 

The role of DnaA protein in controlling DNA synthesis in E. coli is... 

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