Table 1.
Properties of fluorescent proteins in the far-red region of the spectrum.
Fig 1.
A Fluorescence lifetime measurements of mutants from site-directed mutagenesis of mCardinal measured in HEK393T cells by FLIM. B Fluorescence lifetime measurement of mutants from site-directed mutagenesis of mCardinal and mKate2 (in HEK293T cell lysates) and SL72 from the mScarlet/mCardinal shuffle library (in bacterial suspension) measured on the PTI fluorimeter. C Excitation and D Emission spectra of fluorescence proteins with the longest fluorescence lifetimes in HEK293T cell lysates, made by site-directed mutagenesis and SL72 from the mCardinal/mScarlet Shuffle library (measured in bacterial suspension).
Fig 2.
Fluorescence lifetime plotted against peak emission wavelength for all unique fluorescent proteins generated either by site-directed mutagenesis or from the mScarlet/mCardinal shuffle library.
Fig 3.
Residue importance for determining fluorescence lifetime (grey bars) or on peak emission wavelength (black bars) scaled as a proportion of the maximum importance for each property.
Positive numbers indicate that the residue is of relative importance and negative values mean the residue is not important. Numbers on the x-axis correspond to residue number in mCardinal. The secondary structure of the protein is illustrated along the bottom of the graph. Residues 58–79 make up the alpha helix and loop that hold the chromophore (Red pentagon) inside the β-barrel. Numbers highlighted in red indicated those residues on the β-sheet or loop regions that face inside the β-barrel and thus, could interact with the chromophore. Red arrows indicate residues that were independently selected for the site-directed mutagenesis in mCardinal or mKate2.