Flavoproteins fluorescence

B. Chance, B. Schoener, R. Oshino, F. Itshak, and Y. Nakase, Oxidation-reduction ratio studies of mitochondria in freeze-trapped samples. NADII and flavoprotein fluorescence signals, J. Biol. Chem. 254, 4764-4771 (1979). 

W. Halangk and W. S. Kunz, Use of NAD(P)H and flavoprotein fluorescence signals to characterize the redox state ofpyridine nucleotides in epididymal bull spermatozoa, Biochim. Biophys. Acta 1056, 273-278 (1991). 

NADH and oxidized flavin (flavoprotein) fluorescence can be used as an indicator of the redox states of cells challenged with a variety of chemical effectors, and has been used to correlate the oxidative status of cells in situ and in vivo to functions such as electrical activity in brain tissue. The fluorescence of NADH also has been used to characterize the in vitro activity of individual lactate dehydrogenase molecules, which reduce the nonfluorescent NAD+... 

Kunz WS, Gellerich FN. Quantification of the content of fluorescent flavoproteins in mitochondria from liver, kidney cortex, skeletal muscle, and brain. Biochem Med Metab Biol 1993 50 103-110. 

We should note that there are other intrinsic fluoro-phores that can be used in binding studies. NADH and NADPH are both highly fluorescent coenzymes. So too are riboflavin and FAD, but flavoproteins are not typi-... 

Riboflavin (vitamin B2 6.18) consists of an isoalloxazine ring linked to an alcohol derived from ribose. The ribose side chain of riboflavin can be modified by the formation of a phosphoester (forming flavin mononucleotide, FMN, 6.19). FMN can be joined to adenine monophosphate to form flavin adenine dinucleotide (FAD, 6.20). FMN and FAD act as co-enzymes by accepting or donating two hydrogen atoms and thus are involved in redox reactions. Flavoprotein enzymes are involved in many metabolic pathways. Riboflavin is a yellow-green fluorescent compound and, in addition to its role as a vitamin, it is responsible for the colour of milk serum (Chapter 11). 

It is desirable to enhance the accuracy of the method developed for the resolution of intracellular coenzyme fluorescence spectra into their components (free vs. bound reduced pyridine nucleotides (7,8,9,10,11, 28-34), flavoproteins (35,36, 37,38). This resol utionTnay Tie important to understand ancT evaluate different metabolic steady states, drug effects, pathological conditions or divergence between cell types and their transformed variants. 

The fluorescence intensity and anisotropy of the flavin of a flavoprotein have been measured from 0 to 4 kbar. We notice an increase in the fluorescence intensity and a decrease in the fluorescence anisotropy. Explain the results in five to seven lines. 

Since the pioneering work by Cotton et al. on heme proteins (Cotton et al., 1980), surface enhanced resonance Raman spectroscopy (SERRS), Sec. 6.1, has been used to study a large variety of biomolecules, such as retinal proteins (Nabiev et al., 1985), flavoproteins (Coperland et al., 1984 Holt and Cotton, 1987), chlorophylls (Cotton and Van Duyne, 1982 Hildebrandt and Spiro, 1988), and oxyhemoglobins (de Groot and Hesters, 1987). The advantages of this technique include low sample concentration and fluorescence quenching. The main question is whether or not the native structure and function of the molecule is preserved on the metal surface. 

Kindzelskii, A. and Petty, H. R. (2004). Fluorescence spectroscopic detection of mitochondrial flavoprotein redox oscillations and transient reduction of the NADPH oxidase-associated flavoprotein in leukocytes European Biophysics Journal with Biophysics Letters 33 291-299. 

Some proteins contain other native fluorophores in addition to fluorescent amino acids. These include cofactors such as nicotinamide adenine dinucleotide (fluorescent in its reduced, NADH state) and flavin adenine dinucleotide (FAD). NADH is weakly fluorescent in water, but its fluorescence yield increases markedly on binding to a protein-binding site with an emission peak around 470 nm (3). FAD and flavin mononucleotide (FMN) are also fluorescent with an emission maximum around 520 nm, but fluorescence is quenched on binding to many flavoproteins (4). 

Visser AJWG, Ghisla S, Massey V, Muller F, Veeger C. Fluorescence properties of reduced flavins and flavoproteins. Eur. J. Biochem. 1979 101 13-21. 

In this chapter results of the picosecond laser photolysis and transient spectral studies on the photoinduced electron transfer between tryptophan or tyrosine and flavins and the relaxation of the produced ion pair state in some flavoproteins are discussed. Moreover, the dynamics of quenching of tryptophan fluorescence in proteins is discussed on the basis of the equations derived by the present authors talcing into account the internal rotation of excited tryptophan which is undergoing the charge transfer interaction with a nearby quencher or energy transfer to an acceptor in proteins. The results of such studies could also help to understand primary processes of the biological photosynthetic reactions and photoreceptors, since both the photoinduced electron transfer and energy transfer phenomena between chromophores of proteins play essential roles in these systems. 

Fluorescence correlation spectroscopy of flavines and flavoproteins 01MI146. 

Protein fluorescence is also a useful signal in such studies. In some cases, in addition to its aromatic amino acid residues, an enzyme may possess other useful chromophores. Thus rapid reaction studies of haemoproteins and flavoproteins, for example, have relied heavily on the spectral properties of the prosthetic groups of these enzymes. 

Intrinsic fluorescence is relatively rare in biological molecules. Most of the naturally occurring nucleic acids, carbohydrates and lipids show little or no useful fluorescence in the normal UV/visible region. In proteins, fluorescence can only be seen from tryptophan residues and, to a lesser extent, tyrosine side chains. This intrinsic protein fluorescence can be used in a number of practical applications. Typical Trp and protein emission spectra are shown in Figures 2.24 and 2.25. Some proteins contain intrinsically fluorescent prosthetic groups such as reduced pyridine nucleotides and flavoproteins, and the chlorophylls from green plants show a red fluorescence emission. 

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