PHOTOTRANSFORMATION OF THE WILD-TYPE AEQUOREA VICTORIA GREEN FLUORESCENT PROTEIN WITH UV-AND VISIBLE LIGHT LEADS TO DECARBOXYLATION OF GLUTAMATE 222

Phototransformation of Aequorea victoria Green Fluorescent Protein (GFP) was discovered shortly after the cloning of the gfp gene.¹ UV/Vis light transforms the species absorbing at 398 nm (GFP₃₉₈), with a neutral, phenolic chromophore, into an ionic species absorbing at 483 nm (GFP₄₈₃), with a phenolate containing chromophore.^2,3 Besides its role as transient proton acceptor in the excited state, Glu 222 has also been suggested to function as the stable acceptor for the proton that is lost from the chromophore upon photoconversion. Conformational changes such as rotamer reorientation of a threonine side chain (Thr 203) and syn/anti-isomerisation of Glu 222 have been proposed to compete with reformation of the protonated ground state of the chromophore.⁴ However, a direct test of this model with FTIR spectroscopy turned out negative, indicating that Glu 222 is not the stable proton acceptor…