When the wild-type green fluorescent protein (wt GFP) chromophore, p -hydroxybenzylidene-imidazolinone (p -HBDI), was modified into various analogues, its fluorescent state and relaxation pathways were also changed dramatically. In contrast to p -HBDI and its various analogues, such as the o -hydroxy analogue (o -HBDI), the o -sulfonamide analogue (o -TsABDI), the p -sulfonamide analogue (p -TsABDI) and the p -amino analogue (p -ABDI), the p -amido analogue (p -AABDI) of wt GFP chromophore displays single fluorescence and its electronic absorption and fluorescence emission do not involve a charge transfer. Its ground-state N–H acid strength (p K a) is 19.6 in DMSO, which is not very acidic, so, unlike p -TsABDI, p -AABDI does not undergo an excited-state proton transfer. Its S 1 excited state decays mainly along τ torsion through the S 1 /S 0 conical intersection with a barrier of 2.8 kcal/mol. This was confirmed by the cis - trans photoisomerization experiment, in which 47% of cis-p -AABDI was converted to its trans -form right after 20 min irradiation with 350 nm UV light. Image 1 [ABSTRACT FROM AUTHOR]