Resonance energy transfer single-molecule

Although much of the book focuses on physical theory, 1 have emphasized aspects of optical spectroscopy that are especially pertinent to molecular biophysics, and 1 have drawn most of the examples from this area. The book therefore covers topics that receive little attention in most general books on molecular spectroscopy, including exciton interactions, resonance energy transfer, single molecule spectroscopy, high-resolution fluorescence microscopy, femtosecond pump-probe spectroscopy, and photon echoes. It says less than is customary about atomic spectroscopy and about rotational and vibrational spectroscopy of... 

Deniz A A, Dahan M, Grunwell J R, Ha T, Faulhaber A E, Chemla D S, Weiss S and Schultz P G 1999 Single-pair fluorescence resonance energy transfer on freely diffusing molecules observation of Forster distance dependence and subpopulations Proc. Natl Acad. Sc/. USA 96 3670-5... 

Oswald B, Gruber M, Bohmer M, Lehmann F, Probst M, Wolfbeis OS (2001) Novel diode laser-compatible fluorophores and their application to single molecule detection, protein labeling and fluorescence resonance energy transfer immunoassay. Photochem Photobiol 74 237-245... 

Ha, T. (2001). Single-molecule fluorescence resonance energy transfer. Methods 25, 78-86. 

Selvin, P. R., Ha, T., Enderle, T., Ogletree, D. F., Chemla, D. S. and Weiss, S. (1996). Fluorescence resonance energy transfer between a single donor and a single acceptor molecule. Biophys. J. 70, Wp302-Wp302. 

Eggeling, C., Widengren, J., Brand, L., Schaffer, J., Felekyan, S. and Seidel, C. A. (2006). Analysis of photobleaching in single-molecule multicolor excitation and Forster resonance energy transfer measurements. J. Phys. Chem. A 110, 2979-95. 

Scheme 17. Bis-chromophore 54 serves as an example for the detection of fluorescence resonance energy transfer at the single molecule level.

The intersubunit rotation is required for translocation as ribosomes trapped in the nonrotated state by an engineered intersubunit disulfide bridge fail in tRNA-mRNA movement. Real-time observation of intersubunit movement by fluorescence resonance energy transfer (FRET) showed that intersubunit movement occurs concomitantly with hybrid state formation, and that the rotated state can be trapped by the antibiotic viomycin. Similarly to the fluctuation of tRNAs between classical and hybrid states, single-molecule studies have detected spontaneous intersubunit movement where the 3 OS subunit fluctuates between a rotated... 

Abstract. Various ways of extracting information on the conformational structure, dynamics and correlations between them from single-molecule measurements of florescence resonance energy transfer are surveyed. The information obtained via those various ways is then analyzed in detail in the case of an off-lattice model of a two-stranded coiled-coil polypeptide that follows Langevin dynamics. The analysis includes a consideration of the cases of a freely diffusing and surface-immobilized polypeptide as well as the effect of different types of surface and denaturation conditions. 

We have developed the splinted RNA ligation procedure outlined in this chapter to generate site-specifically dye-labeled telomerase RNA constructs. These modified telomerase RNA constructs may be used to characterize dynamic RNA structural properties using Forster resonance energy transfer (FRET) (Stone et al., 2007). Our laboratory specializes in single molecule FRET measurements, which facilitates the direct observation of transient RNA structural states. The details of single molecule FRET... 

Fluorescence resonance energy transfer (FRET) is a spectroscopic means of obtaining distance information over a range up to 80 A in solution. It is based on the dipolar coupling between the electronic transition moments of a donor and acceptor fluorophore attached at known positions on the RNA species of interest. It can be applied in ensembles of molecules, either by steady-state fluorescence or by lifetime measurements, but it is also very appropriate for single-molecule studies. In addition to the provision of distance information, recent studies have emphasized the orientation dependence of energy transfer. 

Fluorescence-based detection methods are the most commonly used readouts for HTS as these readouts are sensitive, usually homogeneous and can be readily miniaturised, even down to the single molecule level.7,8 Fluorescent signals can be detected by methods such as fluorescence intensity (FI), fluorescence polarisation (FP) or anisotropy (FA), fluorescence resonance energy transfer (FRET), time-resolved fluorescence resonance energy transfer (TR-FRET) and fluorescence intensity life time (FLIM). Confocal single molecule techniques such as fluorescence correlation spectroscopy (FCS) and one- or two-dimensional fluorescence intensity distribution analysis (ID FID A, 2D FIDA) have been reported but are not commonly used. 

Ha T. Single-molecule fluorescence resonance energy transfer. 

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