Structural analysis of modified NCP using mass spectrometry

2020 Fiscal Year Final Research Report

• Project/Area Number: 17K07313
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Yokohama City University
• Principal Investigator: AKASHI Satoko 横浜市立大学, 生命医科学研究科, 教授 (10280728)
• Project Period (FY): 2017-04-01 – 2021-03-31
• Keywords: 質量分析 / ヌクレオソームコア粒子 / 構造解析

Outline of Final Research Achievements

To clarify structural changes in nucleosomes caused by modifications,　such　as　DNA　methylation　and　histone　acetylation,　we　performed　biophysical　analysis of modified nucleosome core particles (NCPs) that were reconstituted in vitro. In the analysis of modified NCPs, mono- and di-NCP were prepared with 147 and 366 base-pair (bp) DNA oligomers. Further, we prepared NCPs with DNA oligomers having AT or CG rich sequences, and characterized their biophysical properties by mass spectrometry and small angle X-ray scattering. No structural information is available for these NCPs due to difficulty in crystallization. We established a mass spectrometric method to reliably determine the methylation levels of DNA fragments with ~400 bp.

Free Research Field

構造生物化学

Academic Significance and Societal Importance of the Research Achievements

極めて多くのサイトがメチル化されたDNAを用いて再構成されたDNAの構造解析のための要素技術を質量分析を駆使して開発できたことは、エピゲノム科学研究の基盤構築に貢献できると考えられる。特に、今後クライオ電顕による構造解析を組み合わせることで、このような研究が大幅に進展すると考えられるが、そのための準備段階として、有用な知見が得られた。