| RRC ID |
81679
|
| 著者 |
Hashizume R, Fujii H, Mehta S, Ota K, Qian Y, Zhu W, Drobizhev M, Nasu Y, Zhang J, Bito H, Campbell RE.
|
| タイトル |
A genetically encoded far-red fluorescent calcium ion biosensor derived from a biliverdin-binding protein.
|
| ジャーナル |
Protein Sci
|
| Abstract |
Far-red and near-infrared (NIR) genetically encoded calcium ion (Ca2+ ) indicators (GECIs) are powerful tools for in vivo and multiplexed imaging of neural activity and cell signaling. Inspired by a previous report to engineer a far-red fluorescent protein (FP) from a biliverdin (BV)-binding NIR FP, we have developed a far-red fluorescent GECI, designated iBB-GECO1, from a previously reported NIR GECI. iBB-GECO1 exhibits a relatively high molecular brightness, an inverse response to Ca2+ with ΔF/Fmin = -13, and a near-optimal dissociation constant (Kd ) for Ca2+ of 105 nM. We demonstrate the utility of iBB-GECO1 for four-color multiplexed imaging in MIN6 cells and five-color imaging in HEK293T cells. Like other BV-binding GECIs, iBB-GECO1 did not give robust signals during in vivo imaging of neural activity in mice, but did provide promising results that will guide future engineering efforts. SIGNIFICANCE: Genetically encoded calcium ion (Ca2+ ) indicators (GECIs) compatible with common far-red laser lines (~630-640 nm) on commercial microscopes are of critical importance for their widespread application to deep-tissue multiplexed imaging of neural activity. In this study, we engineered a far-red excitable fluorescent GECI, designated iBB-GECO1, that exhibits a range of preferable specifications such as high brightness, large fluorescence response to Ca2+ , and compatibility with multiplexed imaging in mammalian cells.
|
| 巻・号 |
31(10)
|
| ページ |
e4440
|
| 公開日 |
2022-10-1
|
| DOI |
10.1002/pro.4440
|
| PMID |
36173169
|
| PMC |
PMC9518226
|
| MeSH |
Animals
Biliverdine* / metabolism
Biosensing Techniques*
Calcium / metabolism
Carrier Proteins
HEK293 Cells
Humans
Ions
Luminescent Proteins / chemistry
Mice
|
| IF |
3.876
|
| リソース情報 |
| 遺伝子材料 |
pCAG-iBB-GECO1 (RDB20303)
pCaMKII-iBB-GECO1 (RDB20304)
pCAGGS (RDB08938) |
