Fluorophores quenchers, structures

After scission of the mutant DNA, the FokFDNA assembly remains intact as a FokI cutter unit, which can hybridize to a signaling hairpin containing a fluorophore-quencher pair at its end. This hairpin structure acts as the fuel for the DNA machine. After hybridization, the FokI cutter unit cleaves the hybridized fuel molecule, and upon its spontaneous release the waste product diffuses away. 

The high photostability of the nanoparticle originates from protection provided by the silica matrix for the embedded fluorophores. Environmental oxygen is a universal quencher for fluorophores in aqueous solution and the network structure of the silica matrix reduces diffusion of environmental oxygen to the fluorophores. Thus, the photostability of embedded fluorophores is greatly improved [3]. 

When a protein possesses two or several Trp residues, when quenchers such as iodide, cesium, or acrylamide are used, and if all Trp residues are not accessible to the quencher, the Stern-Volmer equation yields a downward curvature. In this case, we have selective quenching (Figure 10.5b). From the linear part of the plot, we can calculate the value of the Stern-Volmer constant corresponding to the interaction between the quencher and accessible Trp residues. Upon complete denaturation and loss of the tertiary structure of a protein, all Trp residues will be accessible to the quencher. In this case, the Stern-Volmer plot will show an upward curvature. In summary, inhibition of the protein fluorescence with two or several Trp residues can yield three different representations for the Stern-Volmer equations, depending on the accessibility of the fluorophore to the quencher. 

Molecular beacons (MB) are stem-loop hairpin oligonucleotide structures that have a fluorescent dye at one end and a fluorescence quencher at the other. In the hairpin state, the quencher and fluorophore are in close proximity and therefore there is no fluorescence from the probe. However, when the MB binds to a complementary oligonucleotide as a duplex then the fluorophore and quencher are separated and the fluorophore can emit fluorescence. They are particularly useful in monitoring reactions with time, e.g., in PCR, " rolling circle amplification, hybridisation, telomerase activity, ligation... 

Figure 36 The structure-switching signaling aptamer. A DNA duplex composed of three strands of DNA places a fluorophore (F) close to a quencher group (Q). Upon addition of thrombin, the QDNA piece is released, and the fluorescence increases (See ref 67).

Where kp and km are the rate constant of fluorescence and non-radiative processes, respectively. The fluorescence quantum yield (Of) value in the range of 0.0 to 1.0. If the non-radiative relaxation is fast compared to fluorescence (km > k,), O will be small, that is the compound will fluoresce very little or not at all. Often different non-radiative events are limited in the solid phase, and long-lived luminescence (e.g. phosphorescence) is often studied in frozen solution or other solid phases. Quenchers make non-radiative relaxation routes more favorable and often there is a simple relation between 0 and the quencher concentration. The hest-known quencher is probably O2, which quench almost all fluorophores other quenchers only quench a limited range of fluorophores. If a molecule is subject to intramolecular quenching, O may yield information about the structure. 

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