Genetically encoded biosensors based on Förster
resonance
energy transfer (FRET) are indispensable tools for monitoring biochemical
changes in cells. Green and red fluorescent protein-based FRET pairs
offer advantages over the classically employed cyan and yellow fluorescent
protein pairs, such as better spectral separation, lower phototoxicity,
and less autofluorescence. Here, we describe the development of an
mScarlet-derived green fluorescent protein (designated as mWatermelon)
and its use as a FRET donor to the red fluorescent protein mScarlet-I
as a FRET acceptor. We tested the functionality of this FRET pair
by engineering biosensors for the detection of protease activity,
Ca2+, and K+. Furthermore, we described a strategy
to enhance the FRET efficiency of these biosensors by modulating the
intramolecular association between mWatermelon and mScarlet-I.
Genetically encoded biosensors based on Förster resonance energy transfer (FRET) are indispensable tools for monitoring biochemical changes in cells. Green and red fluorescent protein-based FRET pairs offer advantages over the classically employed cyan and yellow fluorescent protein pairs, such as better spectral separation, lower phototoxicity, and less autofluorescence. Here, we describe the development of an mScarlet-derived green fluorescent protein (designated as mWatermelon) and its use as a FRET donor to the red fluorescent protein mScarlet-I as a FRET acceptor. We tested the functionality of this FRET pair by engineering biosensors for the detection of protease activity, Ca2+, and K+. Furthermore, we described a strategy to enhance the FRET efficiency of these biosensors by modulating the intramolecular association between mWatermelon and mScarlet-I.
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