Green and Red Fluorescent Proteins: Photo- and Thermally Induced Dynamics Probed by Site-Selective Spectroscopy and Hole Burning

• Published in: Chemphyschem Vol. 6; no. 5; pp. 838 - 849
• Main Authors: Bonsma, S., Purchase, R., Jezowski, S., Gallus, J., Könz, F., Völker, S.
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 13.05.2005; WILEY‐VCH Verlag; Wiley
• Subjects: Analytical, structural and metabolic biochemistry; Biological and medical sciences; Biophysics - methods; Chemistry, Physical - methods; Electrons; energy transfer; Fluorescence Resonance Energy Transfer - methods; fluorescence spectroscopy; Fundamental and applied biological sciences. Psychology; Green Fluorescent Proteins - chemistry; Green Fluorescent Proteins - metabolism; Hot Temperature; laser spectroscopy; Lasers; Light; Luminescent Proteins - chemistry; Luminescent Proteins - pharmacology; Microscopy, Fluorescence; Models, Chemical; Molecular Conformation; Mutation; photochemistry; Protein Conformation; Protein Structure, Secondary; Proteins; Red Fluorescent Protein; Spectrophotometry - methods; Temperature; High resolution; Label; Molecular interaction; Fluorescence spectrometry; Design; Resonance fluorescence; Signal; Dynamics; Imaging; Light; Optical properties; proteins; Molecular cloning; Interaction energy; Low temperature; Photochemistry; Use; fluorescence spectroscopy; laser spectroscopy; Imagery; Complexes; Electronic structure; Laser; Green fluorescent protein; Mutation; Technique; Energy transfer
• ISSN: 1439-4235; 1439-7641
• DOI: 10.1002/cphc.200500005

The cloning and expression of autofluorescent proteins in living matter, combined with modern imaging techniques, have thoroughly changed the world of bioscience. In particular, such proteins are widely used as genetically encoded labels to track the movement of proteins as reporters of cellular signals and to study protein–protein interactions by fluorescence resonance energy transfer (FRET). Their optical properties, however, are complex and it is important to understand these for the correct interpretation of imaging data and for the design of new fluorescent mutants. In this Minireview we start with a short survey of the field and then focus on the photo‐ and thermally induced dynamics of green and red fluorescent proteins. In particular, we show how fluorescence line narrowing and high‐resolution spectral hole burning at low temperatures can be used to unravel the photophysics and photochemistry and shed light on the intricate electronic structure of these proteins. Green and red fluorescent proteins are favorite markers for tracking motions and protein–protein interactions in live cells. This Minireview surveys the elusive photophysics of green wt‐GFP and red DsRed (see picture) by site‐selective spectroscopy and hole burning at liquid‐helium temperature and examines the changes that occur upon raising the temperature. The physical implications of the observed photoconversions, energy‐transfer and dephasing processes are discussed.