Far-red fluorescent proteins

Gurskaya NG, Fradkov AF, Terskikh A, Matz MV, Labas YA, Martynov VI, Yanushevich YG, Lukyanov KA, Lukyanov SA (2001) GFP-like chromoproteins as a source of far-red fluorescent proteins. FEBS Lett 507 16-20... 

Wilmann PG, Petersen J, Pettikiriarachchi A, Buckle AM, Smith SC, Olsen S, Perugini MA, Devenish RJ, Prescott M, Rossjohn J (2005) The 2.1 angstrom crystal structure of the far-red fluorescent protein HcRed Inherent conformational flexibility of the chromophore. J Mol Biol 349 223-237... 

Petersen J, Wilmann PG, Beddoe T, Oakley AJ, Devenish RJ, Prescott M, Rossjohn J (2003) The 2.0-A crystal structure of eqFP611, a far red fluorescent protein from the sea anemone Entacmaea quadricolor. J Biol Chem 278 44626-44631... 

Pletnev S, Shcherbo D, Chudakov DM, Pletneva N, Merzlyak EM, Wlodawer A, Dauter Z, Pletnev V (2008) A crystallographic study of bright far-red fluorescent protein mKate reveals pH-induced cis-trans Isomerization of the chromophore. J Biol Chem 283 28980-28987... 

TurboFP was also used as a basis for far-red fluorescent proteins (fRFPs). Residues surrounding the chromophore were mutagenized to create a library, which was subsequently subjected to random mutagenesis. A bright far-red variant with excitation and emission maxima at 588 and 635 nm, respectively was isolated and named Katushka [79]. This fast-maturing protein has an... 

Tsai TH, Lin CY, Tsai HJ, Chen SY, Tai SP, Lin KH. Sun CK (2006) Biomolecular imaging based on far-red fluorescent protein with a high two-photon excitation action cross section. Opt Lett 31 930-932... 

Typical spectra for the known red and far-red fluorescent proteins are given in Fig. (16). 

Fradkov, A. F. (2000). Novel fluorescent protein from Discosoma coral and its mutants possesses a unique far-red fluorescence. FEBS Lett. 479 127-130. 

Shu X, Wang L, Colip L, Kallio K, Remington SJ (2009) Unique interactions between the chromophore and glutamate 16 lead to far-red emission in a red fluorescent protein. Protein Sci 18 460 166... 

Mutants of monomeric red fluorescent protein mRFPl at residue 66 structure modeling by molecular dynamics and search for correlations with spectral properties. Biochemistry, Vol. 73, No. 10, pp. 1082-1095, ISSN 0006-2979 Kredel, S. Nienhaus, K Oswald, F. Wolff, M. Ivanchenko, S. Cymer, F. Jeromin, A. Michel, F. J. Spindler, K.D. Heilker, R. Nienhaus, G.U Wiedenmann, J. (2008). Optimized and far-red-emitting variants of fluorescent protein eqFP611. Chem. Biol, Vol. 15, pp. 224-233, ISSN 1074-5521... 

Stiel AC, Andresen M, Bock H et al (2008) Generation of monomeric reversibly switchable red fluorescent proteins for far-field fluorescence nanoscopy. Biophys J 95 2989-2997... 

Direct labeling of a biomolecule involves the introduction of a covalently linked fluorophore in the nucleic acid sequence or in the amino acid sequence of a protein or antibody. Fluorescein, rhodamine derivatives, the Alexa, and BODIPY dyes (Molecular Probes [92]) as well as the cyanine dyes (Amersham Biosciences [134]) are widely used labels. These probe families show different absorption and emission wavelengths and span the whole visible spectrum (e.g., Alexa Fluor dyes show UV excitation at 350 nm to far red excitation at 633 nm). Furthermore, for differential expression analysis, probe families with similar chemical structures but different spectroscopic properties are desirable, for example the cyanine dyes Cy3 and Cy5 (excitation at 548 and 646 nm, respectively). The design of fluorescent labels is still an active area of research, and various new dyes have been reported that differ in terms of decay times, wavelength, conjugatibility, and quantum yields before and after conjugation [135]. New ruthenium markers have been reported as well [136]. 

In 1962, Olson and Romano isolated and purified a water-soluble BChl a-protein from green bacteria. Soon afterwards, the crystalline form of the protein was also obtained . It was in fact the first photosynthetic pigment-protein to be crystallized. Chemical studies showed the BChl a-protein to have a molecular weight of 150 kDa and to contain 21 BChl a molecules. It has a major absorption band in the far-red at 809 nm, which is ascribed to the Qy transition of BChl a, and a corresponding fluorescence band at 818 nm. Its circular-dichroism spectrum in the far-red absorption-band region shows multiple components which can be interpreted in terms ofexciton interactions between the BChl a molecules " . Fluorescence studies of the BChl a-protein in situ indicate that it functions as an intermediate in exd-tation-energy transfer from BChl c in chlorosomes to the BChl-dimer reaction center, P840 ... 

In 2002 Labas and co-workers investigated numerous Anthozoan species and were able to clone 11 new fluorescent proteins with colors ranging from green to red [34]. An additional protein (eqFP611) emitting far-red light was isolated and cloned from Entacmaea quadricolor another anthozoan species [35]. 

Set the excitation wavelength at 325 nm (the red edlge of the eADP absorption spectrum). Excitation at 325 ran. far from the protein absorption maximum, will eliminate, to a great extent, the correction for the bacl pnund of the protein fluorescence. Set the emission wavelength at 410 nm (the maximum of the eADP emission spectrum). 

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