Portal de Periódicos da CAPES

[32] Spectral variants of green fluorescent protein

1999; Academic Press; Linguagem: Inglês

10.1016/s0076-6879(99)02034-0

1557-7988

Gottfried J. Palm, Alexander Wlodawer,

bioluminescence and chemiluminescence research

The use of green fluorescent protein (GFP) for many applications in biological sciences strongly depends on its spectral properties. GFP originates from the jellyfish Aequorea victoria, in which its fluorescence serves to convert the blue light produced by aequorin to green light. Purified protein that retains fluorescence has been the object of intense investigations for many years. Denaturation by harsh treatment, however, can abolish fluorescence and change the absorption spectrum of GFP. The spectrum of the native protein is also affected by many other parameters, such as mutations, protein concentration, ionic strength, temperature, and pH. However, the stability of GFP in a wide pH range (pH 5-12), at high temperature (Tm = 78°), in chaotropic reagents (8 M urea), and against proteolysis makes it attractive for practical use. Expression has been optimized by mutations that change the promoter, ribosome binding site, or codon usage; eliminate splicing sites; or enhance folding. Even though such procedures can drastically increase the overall fluorescence by GFP per cell, as the spectra of purified GFP remain the same in all these cases. Numerous mutations in and around the chromophore, however, do change the spectra. Taken together, these advances enable the use of GFP for detection of promoter activity and for localization of fusion proteins. New applications such as simultaneous localization of two fusion proteins and fluorescence resonance energy transfer (FRET) for detection of Ca2+ concentration or proteolytic activity have become possible through the use of GFP mutants of different colors.