Structural analysis of the supramolecular complexes involved in DNA replication

2014 Fiscal Year Final Research Report

• Project/Area Number: 24770157
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Biophysics
• Research Institution: National Institute of Genetics
• Principal Investigator: HIZUME Kohji 国立遺伝学研究所, 細胞遺伝研究系, 助教 (10378846)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Keywords: DNA replication / nucleosome / atomic force microscopy

Outline of Final Research Achievements

To complete correctly the replication of the genomic DNA, the initiation of DNA replication is highly regulated by the sequential association of protein complexes on replication origin.
In this study, we assembled these protein complexes on the DNA in vitro and formed relatively huge supramolecular complexes. Both molecular imaging using atomic force microscopy (AFM) and biochemical assays show that nucleosome formation on the DNA containing specific sequence of origin induces stable association of ORC on origin. We also found that nucleosome positioning around origin is rearranged by the addition of ORC.
we also performed observation of MCM-DNA complex by AFM and detected that MCM complex bound to more than two DNA strands was detected, which formed DNA-loop. This result suggests existence of the mechanism of that MCM interacts with higher-order structure of DNA such as DNA-loop rather than that MCM is simply loaded on DNA besides ORC.

Free Research Field

分子生物学