2016 Fiscal Year Final Research Report
Biochemical studies on the replication fork model with three DNA polymerases in human cells
| Project/Area Number |
25440011
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Molecular biology
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| Research Institution | Kyushu University |
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Principal Investigator |
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| Project Period (FY) |
2013-04-01 – 2017-03-31
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| Keywords | 複製フォーク / DNAポリメラーゼ / in vitro複製 |
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| Outline of Final Research Achievements |
This research aimed to study the mechanism to proceed a replication fork with three functionally distinct DNA polymerases in eukaryotes. I especially focused on a biochemical background to distinguish DNA polymerases for syntheses of leading and lagging strands. Firstly, I reconstituted the replicative DNA helicase, human CMG complex and obtained the highly purified active CMG helicase. Secondly, I analyzed a functional significance of the interaction between the second PCNA loader Ctf18-RFC and DNA polymerase ε. I elucidated that Ctf18-RFC actively loads PCNA only when it complexes with DNA polymerase ε. This result indicated that Ctf18-RFC functions as a component of the leading-strand DNA polymerase complex and actively loads PCNA on the leading strands. Considering the role of RFC for loading of PCNA on the lagging strands, these two PCNA loaders will coordinate PCNA loading on both strands and play a role to distinguish DNA polymerases for syntheses of the two DNA strands.
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| Free Research Field |
分子生物学、生化学
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