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SPECTROPHOTOMETRIC IDENTITY OF THE ENERGY TRANSFER CHROMOPHORES IN RENILLA AND AEQUOREA GREEN‐FLUORESCENT PROTEINS
Author(s) -
Ward William W.,
Cody Chris W.,
Hart Russell C.,
Cormier Milton J.
Publication year - 1980
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1980.tb03755.x
Subject(s) - aequorea victoria , fluorescence , chromophore , pronase , chemistry , green fluorescent protein , denaturation (fissile materials) , guanidine , kilodalton , titration , biochemistry , trypsin , photochemistry , nuclear chemistry , organic chemistry , physics , quantum mechanics , gene , enzyme
— Spectral properties of guanidine‐denaturated and pronase‐digested green‐fluorescent proteins (GFP) from two species of bioluminescent coelenterates have been investigated. Spectrophotometric titrations of Renilla and Aequorea GFP, following denaturation in 6 M guanidine HCl at elevated temperature, revealed identical absorption peaks in acid (383–384 nm) and in alkali (447–448 nm) and a single isosbestic point in the visible region at 405 nm. Both proteins exhibited a spectrophotometric pK. of 8.1 in guanidine ‐HCl. Pronase digestion of the heat‐denaturated GFP's generated a methanol‐soluble blue‐fluorescent peptide with identical fluorescence emission spectra (λ max = 430 nm, uncorrected; φ f1 = 0.003) for both coelenterate species. These data suggest that the large absorption differences between native Renilla and Aequorea GFP molecules result from unique protein environments imported to a common chromophore.