論文 - 詳細
| RRC ID | 84967 |
|---|---|
| 著者 | Laskaratou D, Fernández GS, Coucke Q, Fron E, Rocha S, Hofkens J, Hendrix J, Mizuno H. |
| タイトル | Quantification of FRET-induced angular displacement by monitoring sensitized acceptor anisotropy using a dim fluorescent donor. |
| ジャーナル | Nat Commun |
| Abstract |
Förster resonance energy transfer (FRET) between fluorescent proteins has become a common platform for designing genetically encoded biosensors. For live cell imaging, the acceptor-to-donor intensity ratio is most commonly used to readout FRET efficiency, which largely depends on the proximity between donor and acceptor. Here, we introduce an anisotropy-based mode of FRET detection (FADED: FRET-induced Angular Displacement Evaluation via Dim donor), which probes for relative orientation rather than proximity alteration. A key element in this technique is suppression of donor bleed-through, which allows measuring purer sensitized acceptor anisotropy. This is achieved by developing Geuda Sapphire, a low-quantum-yield FRET-competent fluorescent protein donor. As a proof of principle, Ca2+ sensors were designed using calmodulin as a sensing domain, showing sigmoidal dose response to Ca2+. By monitoring the anisotropy, a Ca2+ rise in living HeLa cells is observed upon histamine challenging. We conclude that FADED provides a method for quantifying the angular displacement via FRET. |
| 巻・号 | 12(1) |
| ページ | 2541 |
| 公開日 | 2021-5-5 |
| DOI | 10.1038/s41467-021-22816-7 |
| PII | 10.1038/s41467-021-22816-7 |
| PMID | 33953187 |
| PMC | PMC8099864 |
| MeSH | Anisotropy Bacterial Proteins / metabolism Biosensing Techniques Escherichia coli / genetics Escherichia coli / metabolism Fluorescence Resonance Energy Transfer / methods* HeLa Cells Humans Optical Imaging / methods* |
| IF | 12.121 |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | X(Twitter) |
| 各媒体での言及数の合計 | 23 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 0.0 |
| リソース情報 | |
| 遺伝子材料 | pcDNA3-GeuSapVC2.60 (RDB20914) pcDNA3-GeuSapVC3.60 (RDB20915) pRSET-GeudaSapphire (RDB20916) pRSET-GeuSV11.5 (RDB20917) |