2015/01/08

Novel mechanism of DNA duplex unwinding by a single protein

超分子構造研究室・白木原研究室

Structural basis for replication origin unwinding by an initiator-primase of plasmid ColE2-P9: Duplex DNA unwinding by a single protein

Hiroshi Itou, Masaru Yagura, Yasuo Shirakihara, and Tateo Itoh
Journal of Biological Chemistry, 2015 Feb 6;290(6):3601-3611. DOI: 10.1074/jbc.M114.595645

Duplex DNA is generally unwound by protein oligomers prior to replication. The Rep protein of plasmid ColE2-P9 is an essential initiator for plasmid DNA replication. This protein binds the replication origin (Ori) in a sequence specific manner as a monomer and unwinds DNA. We present the crystal structure of the DNA-binding domain of Rep (E2Rep-DBD) in complex with Ori DNA. The structure unveils the basis for Ori specific recognition by the E2Rep-DBD and also reveals that it unwinds DNA by the concerted actions of its three contiguous structural modules. The structure also shows that the functionally unknown PriCT domain, which forms a compact module, plays a central role in DNA unwinding. The conservation of the PriCT domain in the C-termini of some archaeo-eukaryotic primases, indicates that it likely plays a similar role in these proteins. Thus, this is the first report providing the structural basis for the functional importance of the conserved PriCT domain and also reveals a novel mechanism for DNA unwinding by a single protein.

Figure1

(A) The ribbon model of the complex consists of E2Rep-DBD and Ori DNA. Elongated fold of E2Rep-DBD enables binding specificity and affinity enough to unwind duplex DNA.
(B) Model for unwinding of the complete Ori by Rep. The PriCT-module plays a central role in unwinding of duplex DNA and stabilizing the single-stranded DNA.


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