(2019). , , . doi: 10.1101/677344. Article
|Protein||% Normal Cells||OSER/NE ratio||Cell Type||Temp (˚C)|
|mAvicFP1||66.0 (517 cells)||2.31 ± 0.915 (517 cells)||U2-OS||-|
AausFP4 is the first natural example of Dreiklang-type photochromism, and may help generate other useful variations on this mechanism.
As an apparently superior scaffold to avGFP in many ways, mAvicFP1 may be quickly adaptable to existing probes and biosensors
When expressed in total darkness, AvicFP2 is weakly green fluorescent with spectra suggesting an avGFP-type chromophore. After blue light exposure, AvicFP2 rapidly converts (within seconds to minutes), to a purple-blue CP (peak absorbance at 588 nm) which is completely non-fluorescent.
AvicFP3 is highly homologous to AvicFP2 (96% amino acid identity), and is similarly green fluorescent when expressed and purified in the dark. Like AvicFP2, AvicFP3 converts to a green-absorbing chromoprotein when exposed to blue light. We suspect that AvicFP3 is so highly sensitive to blue light exposure that the act of taking an absorbance measurement is enough to partially photoconvert the protein.
We were surprised to discover a second green-emitting FP in A. cf. australis, AausFP1, that shares only 53% amino acid identity with avGFP (see Fig. 4). AausFP1 is the brightest FP discovered to date, with a nearly perfect quantum yield (0.97) and a peak extinction coefficient of 170,000 M−1cm−1, making it nearly fivefold brighter than EGFP on a per-molecule basis. These already extraordinary properties are further bolstered by a low fluorescence pKa (4.4) and unusually narrow excitation and emission peaks (see Fig. 3; the emission peak of AausFP1 has a full width at half maximum (FWHM) of 19nm, compared to 32nm for EGFP).