2023 Fiscal Year Annual Research Report
Next Generation Biosensors Enabled by High-speed Visualization of Dynamic Mechanisms
Publicly Offered Research
| Project Area | Non-equilibrium-state molecular movies and their applications |
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| Project/Area Number |
22H04743
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| Research Institution | The University of Tokyo |
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Principal Investigator |
キャンベル ロバート.アール 東京大学, 大学院理学系研究科(理学部), 教授 (40831318)
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| Project Period (FY) |
2022-04-01 – 2024-03-31
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| Keywords | 高速分子動画 / 蛍光タンパク質 / イメージング / バイオセンサー / ケージド化合物 |
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| Outline of Annual Research Achievements |
The original aim of this project was to obtain mechanistic insight into the dynamic response of genetically encoded biosensors. Working towards this goal, we have been building upon our expertise in biosensor engineering, chemical biology, and photochemistry to reveal the mechanism of lactate biosensors developed in our lab. In FY2023 our primary focus was on the characterization and application of a photocaged lactate (MNI-L-lac). This work has been made publicly available since January 2024, when we posted a preprint posted to bioRxiv. This manuscript described the synthesis, in vitro characterization, and cell-based characterization of the photocaged MNI-L-lac. This manuscript will be submitted to a peer reviewed journal as soon as a few additional cell-based imaging experiments are completed.
With MNI-L-lac in hand, we are now well positioned to undertake the rapid uncaging of lactate in crystals in order to perform time-lapse structural studies of the biosensor response. Unfortunately, we have continued to struggle to produce diffracting crystals of the apo state. Work towards this goal is continuing in collaboration with the Nureki lab. In the meantime, we have succeeded in obtaining a cryo-EM structure of a lactate-bound lactate biosensor. While this structure can not reveal the full dynamic mechanism that we were hoping for, it does provide important insights into the key interactions chromophore-protein interactions responsible for the fluorescence response. Further efforts to interpret and validate the mechanistic insights are ongoing.
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| Research Progress Status |
令和5年度が最終年度であるため、記入しない。
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| Strategy for Future Research Activity |
令和5年度が最終年度であるため、記入しない。
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