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Nontrivial Effect of the Color-Exchange of a Donor/Acceptor Pair in the Engineering of Förster Resonance Energy Transfer (FRET)-Based Indicators
- Source :
- ACS Chemical Biology. 11:1816-1822
- Publication Year :
- 2016
- Publisher :
- American Chemical Society (ACS), 2016.
-
Abstract
- Genetically encoded indicators driven by the Förster resonance energy transfer (FRET) mechanism are reliable tools for live imaging. While the properties of FRET-based indicators have been improved over the years, they often suffer from a poor dynamic range due to the lack of comprehensive understanding about how to apply an appropriate strategy to optimize the FRET parameters. One of the most successful optimizations is the incorporation of circularly permuted fluorescent proteins (cpFPs). To better understand the effects of this strategy, we systematically investigated the properties of the indicators by utilizing a set of FRET backbones consisting of native or one of the most effective cp variants (cp173FPs) with considerations of their order. As a result, the ordering of donor and acceptor FPs, which has been ignored in previous studies, was found to significantly affect the dynamic range of indicators. By utilizing these backbones, we succeeded in improving a cGMP indicator with 3.6-fold increased dynamic range and in generating an ultrasensitive cAMP indicator capable of environmental imaging, demonstrating the practical importance of the ordering of donors and acceptors in the engineering of FRET-based indicators.
- Subjects :
- 0301 basic medicine
030102 biochemistry & molecular biology
Chemistry
Color
Nanotechnology
General Medicine
Biochemistry
Acceptor
03 medical and health sciences
030104 developmental biology
Förster resonance energy transfer
Fluorescence Resonance Energy Transfer
Molecular Medicine
Biological system
Donor acceptor
Fluorescent Dyes
Subjects
Details
- ISSN :
- 15548937 and 15548929
- Volume :
- 11
- Database :
- OpenAIRE
- Journal :
- ACS Chemical Biology
- Accession number :
- edsair.doi.dedup.....d1b12c8ca12fc35367315d7c76e522ee