2020 Fiscal Year Annual Research Report
Comprehensive analysis of recombination auxiliary factors
| Project/Area Number |
20K15713
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| Research Institution | Tokyo Institute of Technology |
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Principal Investigator |
Argunhan Bilge 東京工業大学, 科学技術創成研究院, 特任助教 (30792759)
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| Project Period (FY) |
2020-04-01 – 2021-03-31
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| Keywords | 相同組換 / DNA二重鎖切断 / DSB 修復 / Rad51 / タンパク質相互作用 / Swi5-Sfr1 / Rad52 / Rad55-Rad57 |
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| Outline of Annual Research Achievements |
DNA double-strand breaks (DSBs) are the severest form of DNA damage. Homologous recombination (HR) is a mechanism that utilizes identical genetic information located elsewhere in the genome as a template for accurate DSB repair. Rad51 is the central protein in HR, but multiple auxiliary factors are involved in modulating Rad51, including Rad52, Rad54, Rad55-Rad57, and Swi5-Sfr1. These auxiliary factors all bind to Rad51, and this binding is thought to be critical for their role in Rad51 potentiation. Despite this, little is known about the nature of these interactions, especially with regards to Rad55-Rad57.
We originally tried to biochemically characterize Rad55-Rad57. Although we succeeded in purification, we were unable to characterize the interaction in-depth due to the biochemical intractability of Rad55-Rad57. We therefore employed a different approach. Structural analysis of Rad51 highlighted a solvent-exposed protruding acidic patch (PAP) comprised of three residues: E205, E206, D209. Our genetic analysis demonstrated that the E206A mutation specifically impairs the interaction with Rad55-Rad57. Moreover, mutation of all three residues to Ala (Rad51-EED) abrogated the interaction with Rad52, further impaired the interaction with Rad55-Rad57, and even impaired the interaction with Rad54. These results demonstrate that the PAP, a newly identified motif of Rad51, functions as an interaction hub for multiple auxiliary factors to facilitate the potentiation of Rad51. These results provided critical new insights into the regulation of HR by auxiliary factors.
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