2018 Fiscal Year Final Research Report
Molecular basis underlying cancer cell proliferation and T-cell anergy induction by diacylglycerol kinase alpha
| Project/Area Number |
17K15444
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Multi-year Fund |
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| Research Field |
Biological pharmacy
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| Research Institution | Kyushu University (2018)
Chiba University (2017) |
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Principal Investigator |
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| Research Collaborator |
Sakane Fumio
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| Project Period (FY) |
2017-04-01 – 2019-03-31
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| Keywords | ジアシルグリセロールキナーゼ / 脂質キナーゼ / マルチドメイン蛋白質 / 構造解析 / 昆虫細胞・バキュロウイルス / EFハンド |
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| Outline of Final Research Achievements |
Diacylglycerol kinases (DGKs) modulate the levels of lipid messenger, DG and PA, thus function as a molecular hub for cellular signaling. To date, however, little progress has been made for the structural biology of DGKs including the structural basis for its catalytic function and regulation. In this study, we have produced a full-length DGKα using baculovirus-insect cell expression system. The purified protein was found to be a monomer and enzymatically active, demonstrating that this approach is very promising to produce DGKα for future functional and structural studies. Furthermore, we have determined the first crystal structure of the N-terminal EF-hand motifs (DGKα-EF) in its Ca2+ bound form. Our biochemical and biophysical analyses also reveal that DGKα-EF adopts a protease-susceptible “open” confirmation and cooperative binding of two Ca2+ ions induce substantial conformational changes, likely regulates intramolecular interactions responsible for the activation of DGKα.
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| Free Research Field |
蛋白質科学，構造生物学，生化学
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| Academic Significance and Societal Importance of the Research Achievements |
脂質キナーゼであるDGKに関する構造生物学的理解は乏しく，活性が確認されて以来約60年，DGK触媒領域の構造は不明のままである．特に，がん免疫治療の標的の一つとして近年注視されているDGKαの構造基盤の解明は，構造に指南された阻害剤開発のためにひも必要である．本研究で初めて確率した昆虫細胞によるDGKα試料の大量発現・精製法およびN末端EFハンドドメインの構造決定と構造変化の解析は，これまで遅れていたDGKの構造研究を進展させるものである．
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