Shedding Light on the Dark and Weakly Fluorescent States of Green Fluorescent Proteins

Recent experiments on various similar green fluorescent protein (GFP) mutants at the single-molecule level and in solution provide evidence of previously unknown short- and long-lived "dark" states and of related excited-state decay channels. Here, we present quantum chemical calculations...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1999-05, Vol.96 (11), p.6177-6182
Main Authors: Weber, Wolfgang, Helms, Volkhard, McCammon, J. Andrew, Langhoff, Peter W.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Amino Acid Substitution
Biochemistry
Biological Sciences
Biology
Chromophores
Fluorescence
Green Fluorescent Proteins
Ground state
Isomerism
Isomerization
Isomers
Kinetics
Lead
Luminescent Proteins - chemistry
Luminescent Proteins - metabolism
Models, Chemical
Models, Molecular
Molecules
Mutagenesis, Site-Directed
Photochemistry
Physics
Proteins
Protons
Quantum Theory
Recombinant Proteins - chemistry
Recombinant Proteins - metabolism
Thermodynamics
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Summary:Recent experiments on various similar green fluorescent protein (GFP) mutants at the single-molecule level and in solution provide evidence of previously unknown short- and long-lived "dark" states and of related excited-state decay channels. Here, we present quantum chemical calculations on cis-trans photoisomerization paths of neutral, anionic, and zwitterionic GFP chromophores in their ground and first singlet excited states that explain the observed behaviors from a common perspective. The results suggest that favorable radiationless decay channels can exist for the different protonation states along these isomerizations, which apparently proceed via conical intersections. These channels are suggested to rationalize the observed dramatic reduction of fluorescence in solution. The observed single-molecule fast blinking is attributed to conversions between the fluorescent anionic and the dark zwitterionic forms whereas slow switching is attributed to conversions between the anionic and the neutral forms. The predicted nonadiabatic crossings are seen to rationalize the origins of a variety of experimental observations on a common basis and may have broad implications for photobiophysical mechanisms in GFP.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.96.11.6177