Regulation of DNA clamp reactions involved in DNA recombination

2018 Fiscal Year Final Research Report

• Project/Area Number: 15K16128
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Risk sciences of radiation and chemicals
• Research Institution: Fujita Health University (2018); Sojo University (2015-2017)
• Principal Investigator: Kawai Akito 藤田医科大学, 医学部, 助教 (20435150)
• Project Period (FY): 2015-04-01 – 2019-03-31
• Keywords: タンパク質の結晶化 / X線結晶構造解析

Outline of Final Research Achievements

In this research, we have carried out structural study of heterotetrameric DNA clamp complexed with holliday junction resolvase stoHjc and RecQ-like DNA helicase stoHjm. stoHjc, stoHjm, and heterotetrameric DNA clamp were overexpressed in E. coli and purified over 98% for the crystallization. The stoHjcーheterotetrameric DNA clamp complex was prepared by mixing each protein solution and purified using a gel filtration column chromatography. After performing the crystallization screenings and the modification of the crystallization conditions, we obtained the crystals of the stoHjcーheterotetrameric DNA clamp complex, stoHjcーheterotetrameric DNA clampーholliday junction complex, and stoHjm, and carried out X-ray diffraction experiments using the synchrotron radiation at Photon Factory and SPring-8.

Free Research Field

構造生物学

Academic Significance and Societal Importance of the Research Achievements

本研究ではこれまで不明であった四量体型DNAクランプの機能を解明するため研究を進め、立体構造が未知であるstoHjcー四量体型DNAクランプーホリデイ構造複合体の構造解析へとつながる結晶化条件の発見に至った。今後、さらに研究を進めることで四量体型DNAクランプの機能解明を行い、この四量体型DNAクランプの機能を利用してDNA修復の正確性、効率を向上につながるアイデアを生み出したい。