## 5 The Specific Case of DnaA in the _E. coli_ Cell Cycle  
### DnaA Background  
DnaA, a protein found in many bacteria, is commonly believed to be responsible for the initiation of chromosome replication in bacteria [49, 69, 70, 71]. In the Beginning, however, DnaA was not present and a more fundamental mechanism operated. DnaA would therefore have evolved as a veneer on this mechanism. If so, it may prove possible to interpret DnaA action in the framework of Dualism. This we now attempt.  
DnaA binds with different affinities to sites in the origin of replication, _oriC_, and elsewhere on the chromosome [72, 73]. The synergistic binding to _oriC_ sites results in the unwinding of an AT-rich region and the opening of the double helix that constitutes the first step in initiation [74]. DnaA binds ATP and ADP, can form oligomers, and interacts with lipids. But what controls DnaA? One regulatory mechanism affects the availability of DnaA: substantial proportion of DnaA binds to the _datA_ locus [53].  
Another regulatory mechanism for controlling DnaA depends on the direct activation of DnaA [75] which results from it binding and hydrolysing ATP (only the ATP-DnaA form is active in initiation [76]). Sufficient activation for initiation requires the reactivation of existing ADP-DnaA since _de novo_ synthesis of DnaA is not enough [77]. This activation can result from the binding of DnaA to either the membrane [78] or DnaA-reactivating sequences, DARS1 and DARS2, on the chromosome [52].  
Another factor to take into account in the control over DnaA is that fluorescent DnaA-GFP hybrids bind not only to _oriC_ and _datA_ to give foci or clusters but also elsewhere in the chromosome to give a helix [79, 80, 81]. It should be noted here and elsewhere that fusions with fluorescent probes such as GFP can lead to artifactual localization of the protein under study [82, 83]. It might be argued that such fusions may sometimes increase slightly the degree of a self-association that occurs naturally. It has been shown recently that MreB, rather than forming a helix on its own, forms a proteolipid domain with fluid phospholipids [154] and these patches move independently along a helical path [84, 85]. It may turn out that the distribution of DnaA and MreB are both controlled by helical lipid domains created by transertion [86].  
How can these characteristics of DnaA be accommodated by the Dualism hypothesis in the context of intensity sensing and quantity sensing (see Table 1 for a summary)?  
\begin{table}
\begin{tabular}{|p{142.3 pt}|p{142.3 pt}|} \hline
**Intensity Sensing** & **Quantity Sensing** \\ \hline
**Superhelicity**. Increased superhelicity during growth might release DnaA from DnaA binding at sites on the chromosome [87]. & **Cardiolipin**. CL detaches DnaA from DNA) [87]. CL domains are also important in division [90]. \\
**ATP levels**. If several DnaA are bound to DARS they can be recharged with ATP (ATP\({}^{\pm}\) condenses before ADP\({}^{\pm}\)) [52]. & **Nucleoid Associated Proteins**. NAPs include HU, H-NS, Fis, IHF, StpA and Dps; each binds to 100s of specific sites + every 100 bp. IHF, HU and Dps interact with DnaA [91, 92]. IHF for example also binds to oriC, helps DnaA bind to datA and binds itself to datA (IHF deletion delays initiation) [93]. H-NS is in a hyperstructure [16] and H-NS deletion delays initiation \\ \hline \end{tabular}
\end{table}
Table 1: Characteristics of the DnaA protein (and factors that may affect it) can be classed as suggestive of its role in either intensity sensing or quantity sensing (see text for details).