a.k.a. GFP with acid tolerance and monomeric properties to illuminate soured environments
|State||Ex λ||Em λ||EC (M-1 cm-1)||QY||Brightness||pKa||Maturation (min)||Lifetime (ns)|
|State||t1/2 (s)||Power||Light||Mode||In Cell||Fusion||˚C||Reference|
|On||73.2||3.7 (W/cm2)||Arc-lamp||Widefield||37.0||Shinoda et al. (2018)|
Gamillus was derived from Gamillus0.5 with the following mutations: M1_A2insVSKGEE/A150_G152delinsPHGP/K180T/E226_V232delinsGMDELYK
Gamillus expressed in HeLa cells also exhibited a photochromic decrease in fluorescence to ∼60% or ∼10% of its initial intensity by exciting with 457–487 nm (GFP excitation range, 470 mW/cm2 for 40 s) or 488–512 nm (YFP excitation range, 360 mW/cm2 for 40 s) mercury arc light, respectively. The decreased intensity was recovered by subsequent irradiation with 352–388 nm light (770 mW/cm2 for 10 s). In contrast, such time-dependent fluorescence intensity changes were negligible using 440–480 nm excitation light as it presumably renders photochromic equilibrium of Gamillus dominant in the ionic state.
One of the possible reasons for the negligible photochromism is a much higher on-switching rate, compared with the switching-off rate. In our estimation, the switching-on rate is ~200-fold higher than switching-off with 405 nm and 488 nm laser illumination, at the same power density... To simplify imaging we therefore selected 440–480 nm illumination to limit the photochromic phenomena. Under this excitation wavelength range, the in vitro brightness of Gamillus (calculated as the product of averaged extinction coefficients over 440–480 nm [ɛ440-480] and quantum yield) is 98% of that of EGFP (ɛ440-480 for Gamillus and EGFP were 28.8 and 35.4 mM−1 cm−1, respectively)