NDLI: Fluorescence polarization of green fluorescence protein.

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Fluorescence polarization of green fluorescence protein.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Shribak, Mykhailo Inoué, Shinya Shimomura, Osamu Goda, Makoto Tran, P. T.
Description	Author Affiliation: Inoué S ( Marine Biological Laboratory, 7 MBL Street, Woods Hole, MA 20543-1015, USA.);
Abstract	We report here the striking anisotropy of fluorescence exhibited by crystals of native green fluorescence protein (GFP). The crystals were generated by water dialysis of highly purified GFP obtained from the jellyfish Aequorea. We find that the fluorescence becomes six times brighter when the excitation, or emission, beam is polarized parallel (compared with perpendicular) to the crystal long axis. Thus, the major dipoles of the fluorophores must be oriented very nearly parallel to the crystal long axis. Observed in a polarizing microscope between parallel polars instead of either a polarizer or analyzer alone, the fluorescence polarization ratio rises to an unexpectedly high value of about 30:1, nearly the product of the fluorescence excitation and emission ratios, suggesting a sensitive method for measuring fluorophore orientations, even of a single fluorophore molecule. We have derived equations that accurately describe the relative fluorescence intensities of crystals oriented in various directions, with the polarizer and analyzer arranged in different configurations. The equations yield relative absorption and fluorescence coefficients for the four transition dipoles involved. Finally, we propose a model in which the elongated crystal is made of GFP molecules that are tilted 60 degrees to align the fluorophores parallel to the crystal long axis. The unit layer in the model may well correspond to the arrangement of functional GFP molecules, to which resonant energy is efficiently transmitted from Ca2+-activated aequorin, in the jellyfish photophores.
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	7
Volume Number	99
Language	English
Publisher	National Academy of Sciences
Publisher Date	2002-04-01
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Luminescent Proteins Chemistry Animals Crystallization Fluorescence Polarization Green Fluorescent Proteins Microscopy, Polarization Scyphozoa Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Multidisciplinary
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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