Reaction progress of chromophore biogenesis in green fluorescent protein

The kinetics of hydrogen peroxide evolution during in vitro green fluorescent protein (GFP) maturation is investigated and it is found that the H2O2 coproduct is generated prior to the acquisition of green fluorescence at a stoichiometry of 1:1 (peroxide/chromophore). The molecular mass of an oxidized intermediate is characterized and it is found that, in air-saturated aqueous solutions, a dehydrogenated species is generated.