Based on the capacity of its fluorescence to vanish and reappear, 22Gm3 was renamed “Dronpa,” after “dron,” a ninja term for vanishing, and “pa,” which stands for photoactivation.
We prepared a cDNA library from Pectiniidae, a species of coral that emits faint fluorescence upon irradiation with ultraviolet light. Approximately 300,000 bacterial colonies containing individual cDNA clones were screened for fluorescence. A single clone (22G) encoding a greenish fluorescent protein with highest homology to mcavGFP (Labas et al 2002) (74.6%) was identified.
We found 22Gm3 to possess a distinctive photosensitivity. Strong excitation at around 490 nm appeared to bleach 22Gm3 more efficiently than other fluorescent proteins, and the bleached protein regained its green fluorescence completely with minimal irradiation at around 400 nm. We intermittently measured the absorbance of a solution containing 22Gm3 at pH 7.4 in a cuvette during continuous illumination at 490 ± 10 nm with a 75-W xenon lamp. After a 40-min incubation, nearly all the protein molecules had been converted into the neutral, nonfluorescent state. Illumination at 400 ± 7.5 nm for several minutes reversed the protein to its original fluorescent state. Thus, 22Gm3 has photochromic behavior, because its fluorescence can be switched on and off by using two different wavelengths of light.