Structural and functional analyses of the DNA replication machinery called replisome complex

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DNA replication processes by a macromolecular complex called replisome, which was made of fork-structured DNA and many protein factors. The aim of this research is to understand the molecular mechanism of assembling and reorganizing of the replisome complex for the precise progress of DNA replication. The originarity of our esearch is that we use hyperthermophilic archaea, which provide very useful experimental materials especially appropriate for the structural analyses of the complexes. We characterized the Cdc6/Orc1 protein that specifically binds to the replication origin (oriC) of the chromosome DNA. Using electron microscopic analyses, we observed the structural properties of this proein, and found that this protein converts its structure depending on the nucleotide cofactors. In addition, we characterized Hef, and Hjm proteins, which were discovered from archaeal cells (Pyrococcus furiosus) by ourselves. These proteins works for the repair systems coupled with DNA replication by converting the fork-structure of replicating DNA to the appropriate form for the following recombinational repair process. Dr.Kohda, a collaborator of this research, has been elucidated that the PriA, which may be the functional homolog of Hef in Esherichia coli, has a recognizing ability of the terminus of the nascent DNA in the replication fork, and provided a structural basis of this recognition mechanism.