Principles of Fluorescence Spectroscopy

PRINCIPLES OF FLUORESCENCE SPECTROSCOPY 83
Figure 3.35. Excitation (top) and emission spectra (bottom) of GFP
mutants. Revised from [129].
The fluorophores in GFPs are formed autocatalytically,
and it is not necessary to add a fluorophore or enzymes to
synthesize the fluorophore. All that is needed is the gene or
mRNA that codes for the amino-acid sequence. As a result
organisms that contain the gene or mRNA can express the
fluorescent protein. This possibility is shown in Figure 3.36
for a Xenopus embryo. While in the blastomer stage the
right and left blastomers were injected with mRNA for a
green or red fluorescent coral protein, respectively. One
week later the descendants of the injected blastomer
showed fluorescence from the fluorescent proteins coded
by the mRNAs.
3.8. OTHER FLUORESCENT PROTEINS
3.8.1. Phytofluors: A New Class of
Fluorescent Probes
A new type of fluorescent probe is the so-called "phytofluors."
These fluorescent probes are derived from the phytochromes,
which are light-sensitive proteins present in
photosynthetic organisms. These proteins allow the organisms
to adjust to external light conditions, and are important
in seed germination, flowering, and regulation of plant
growth. Phytochromes typically contain a nonfluorescent
chromophore that interconverts between two stable forms.
Some phytochromes spontaneously form covalent
adducts 137–139 with phycoerythrobilin. The absorption and
Figure 3.36. Color photograph (right) of a Xenopus embryo injected with the mRNA from a green fluorescent coral protein (right side) and a red fluorescent
coral protein (left side). The left side shows a white light photograph. Reprinted from [125].