Structure and Function of the Bacterial Genome. Charles J. Dorman

Structure and Function of the Bacterial Genome - Charles J. Dorman


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Plasmid research has provided important insights into gene regulation mechanisms, including the provision of early examples of the regulatory roles of small RNA molecules.

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      Many of the cis or trans‐acting components of iteron origins, and their architectures, are reminiscent of oriC on the chromosome. The TrfA replication protein of RK2 interacts with, and recruits, the DnaB helicase. The ability of a plasmid replication protein to recruit a host helicase may be a determining factor limiting plasmid host range (Zhong et al. 2005). TrfA also acts with Hda, the inhibitor of DnaA activity, to prevent over‐initiation of RK2 replication (Kim et al. 2003). It has been suggested that TrfA has a motif that is suitable for interaction with the β‐clamp of E. coli DNA Pol III (Kongsuwan et al. 2006). In addition to DnaB, iteron‐based initiation also requires DnaC (in E. coli), the DnaG primase, DNA gyrase, the Pol III holoenzyme, and the SSB, as is also seen at oriC (Section 1.3). Unlike initiation of chromosome replication at oriC, initiation of plasmid DNA replication at iteron origins is ATP‐independent; there is no requirement for the DnaA‐ATP form of DnaA (Konieczny et al. 2014).

      RK2 and its plasmid relatives use a theta model of DNA replication (Figure 1.12). Plasmid ColE1, the backbone for many cloning vectors used in recombinant DNA technology, also uses theta replication, but differs from RK2‐like plasmids in relying on host factors to open the double‐stranded origin and to prime synthesis (Lilly and Camps 2015; Wang et al. 2004). DNA duplex unwinding is driven by transcription of a stable RNA pre‐primer that forms an R‐loop in the ori region of ColE1. This process is driven by negative supercoiling of the plasmid DNA. RNase H then processes the bound RNA to generate the primer RNA that is then extended by Pol I. This marks the beginning of leading strand DNA synthesis. As the newly synthesised DNA strand makes progress through the plasmid DNA duplex, it base pairs with the template to create a D‐loop that recruits PriA. Pol III takes over leading strand synthesis and initiates the synthesis of the lagging strand; the converging replisomes continue moving until they are at or near the termination site terH (Nakasu and Tomizawa 1992). Gaps between the strands are then filled in by Pol I (Troll et al. 2014).


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