2018 Fiscal Year Final Research Report
substrate-induced product-release mechanism of L-PGDS
| Project/Area Number |
16K18868
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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 |
Physical pharmacy
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| Research Institution | Kindai University |
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Principal Investigator |
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Keywords | プロスタグランジンD2 / 等温滴定型熱測定 / 酵素活性測定 / 構造解析 / PGDS |
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| Outline of Final Research Achievements |
Lipocalin-type prostaglandin (PG) D synthase (L-PGDS) belongs to the lipocalin superfamily which consists of transporter proteins for lipophilic ligands in the extracellular space, and is known as the PGD2-synthesizing enzyme responsible for the sleep regulation. The binding affinity and stoichiometry to each ligand were analyzed by isothermal titration calorimetry (ITC), and the binding region for each ligand on L-PGDS was estimated by NMR titration experiment. The ITC results showed that PGD2 and PGF2α bound to L-PGDS with a stoichiometry of 2 to 1 but PGE2 with a stoichiometry of 1 to 1. In addition, the NMR experiments indicated that PGD2 and PGF2α bound to both the catalytic site containing the Cys65 and non-catalytic site of L-PGDS, while PGE2 bound to only the non-catalytic site.
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| Free Research Field |
熱力学、構造生物学
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| Academic Significance and Societal Importance of the Research Achievements |
L-PGDSの酵素反応モデルで、基質・生成物が2分子結合することは全く考慮されてこなかった。本研究では、1）従来のモデルにない新しい概念（2つの結合サイト・生成物放出）を加えた新規酵素反応モデルが構築し、これまで着目されていなかった親和性の低い結合サイトでの基質認識が生成物の産生速度に影響を与えることを示した。2）L-PGDSの活性中心のみをターゲットにした阻害剤の効果が弱い原因を解明し、さらに、新規結合部位における基質や生成物側の認識に重要な部位の同定とタンパク質側の認識に関与するアミノ酸を同定した。これらの情報は、新たに生成物放出メカニズムをターゲットとした新規薬剤の開発につながるだろう。
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