Flavin radical

As mentioned above, Beinert 3) was first to see flavin radicals in neutral solution, while Michaelis and Schwarzenbach 129) could only infer their existence in the largely disproportionated state of the half reduced neutral flavin system. There are several means for shifting equilibrium between oxidized and reduced flavin towards the radical side  

Protonation The neutral radical, FIH, will accept a proton with a pK of about 3. 

Alkylation Introduction of alkyl groups into the flavin nucleus is a complex matter, which will be extensively reviewed in the dihydroflavin section below. 

In the half reduced system, 5-alkylation brings about comproportionation to the radical. Optical as well as EPR comparison of this stabilized radical, 5-RFl (27, 136), with Beinert s unstable HFl (2) leads to the 

Recomproportionation to yield 1-RFl could only be brought about by further removal of the proton from N(5) in l-RFbedH, which requires very strong base, since it is a typical pyrrole NH. Such a strong base first interferes, however, with the equally present 1- or 2a-RFCx by formation of adducts RFIX (cf. below) which decompose irreversibly  

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Flavin radical anion Neutral flavin radical... 

These include the mitochondrial respiratory chain, key enzymes in fatty acid and amino acid oxidation, and the citric acid cycle. Reoxidation of the reduced flavin in oxygenases and mixed-function oxidases proceeds by way of formation of the flavin radical and flavin hydroperoxide, with the intermediate generation of superoxide and perhydroxyl radicals and hydrogen peroxide. Because of this, flavin oxidases make a significant contribution to the total oxidant stress of the body. 

FIGURE 13.6 Whole bacterial-cell EPR. A frozen concentrated suspension of cells from the sulfate-reducing bacterium Desulfovibrio vulgaris gives an EPR spectrum with only a [2Fe-2S]1+ signal and a flavin radical signal, both from adenosine phosphosulfate reductase. 

In the reduction or oxidation of quinone/ quinol systems, free radicals also appear as intermediate steps, but these are less reactive than flavin radicals. Vitamin E, another qui-none-type redox system (see p.l04), even functions as a radical scavenger, by delocalizing unpaired electrons so effectively that they can no longer react with other molecules. 

MAO oxidizes amine substrates also by a one-electron route via the cyclopropylamine radical cation 8 which undergoes ready ring opening to the iminium radical cation 9 [11]. Then capture by a flavin radical, may cause the enzyme inactivation [12]. This mechanism was established by labeling experiments, Eq.(4) [13]. 

Since the initial observation of flavin radical species by Michaelis and coworkers the involvement of flavins in one-electron oxidation-reduction processes in biological systems has occupied the attention of workers in the field of redox enzymology up to the present time. Flavin coenzymes occupy a unique role in biological oxidations in that they are capable of functioning in either one-electron or two-electron transfer reactions. Due to this amphibolic reactivity, they have been termed in a recent review to be at the crossroads of biological redox processes. 

The absorption spectral properties of the neutral and anionic forms are quite different as shown in Fig. 1. Due to the rapid dismutation of flavin radicals to form an equilibrium mixture with the hydroquinone and oxidized forms of the flavin, special procedures must be employed to measure the spectral properties of free flavin radicals. Nearly quantitative amounts of anion radical can be formed in aprotic solvents under basic conditions Alkylation of the N(5) position of the flavin hydroquinone followed by oxidation results in nearly quantitative formation of the... 

The model system studies of Muller et al. have shown that alkylation of the 0(2) and 0(4) positions of the isolloxazine ring results in a flavin radical with similar ESR and absorption spectral properties as the anion flavin radicals although this flavin species has a neutral charge. Whereas binding of the flavin to its site on... 

ENDOR spectroscopy has proven to be a valuable technique to provide information on both free and protein bound flavin radicals. Since flavin radical ESR spectra can be partially saturated at moderate microwave power, ENDOR spectra may be observed as nuclear spin transitions by detection of changes in the partially saturated ESR signal as a function of nuclear radio frequency. The resonance condition for nuclei (when I = Vz) is described by the following equation ... 

Flash photolysis has proven to be a useful technique for investigating the reactivity of flavin radicals. This approach takes advantage of the fact that illumination of flavins in the presence of suitable reducing agents (e.g. EDTA, phenols, indoles) leads to a one-electron reduction of the flavin triplet state Through the... 

Ehrenberg, A. Flavin radical-metal chelates. In Vitamins and hormones, Vol. 28 (Harris, R. S., Munson, P. L., Diezfalusy, E. eds.) pp. 489-503, New York, London, Academic Press 1970... 

There have been many investigations of phosphotransferases by NMR and EPR methods.681,682 One approach is to use paramagnetic ions such as Mn2+, Cu2+, or Cr3+ to induce nuclear relaxation in substrate and coenzyme molecules at active sites of enzymes. Flavin radicals and specifically introduced nitroxide spin labels can serve as well. Paramagnetic ions greatly increase the rate of magnetic relaxation of nearby nuclei (Chapter 3, Section I). Thus, small amounts of... 

A related reaction that is known to proceed through acetyl-TDP is the previously mentioned bacterial pyruvate oxidase. As seen in Fig. 14-2, this enzyme has its own oxidant, FAD, which is ready to accept the two electrons of Eq. 14-22 to produce bound acetyl-TDP. The electrons may be able to jump directly to the FAD, with thiamin and flavin radicals being formed at an intermediate stage.1353 The electron transfers as well as other aspects of oxidative decarboxylation are discussed in Chapter 15, Section C. 

A long-known characteristic of D-amino acid oxidase is its tendency to form charge-transfer complexes with amines, complexes in which a nonbonding electron has been transferred partially to the flavin. Complete electron transfer would yield a flavin radical and a substrate radical which could be intermediates in a free radical mechanism, as discussed in the next section.256... 

Neutral flavin radicals have a blue color (the wavelength of the absorption maximum, A.max, is -560 nm) but either protonation at N-l or dissociation of a proton from N-5 leads to red cation or anion radicals with imax at -477 nm. Both blue and red radicals are... 

If an enzyme binds a flavin radical much more tightly than the fully oxidized or reduced forms, reduction of the flavoprotein will take place in two one-electron steps. In such proteins the values of E° for the two steps may be widely separated. The best known examples are the small, low-potential electron-carrying proteins known as flavodoxins.266 269a These proteins, which carry electrons between pairs of other redox proteins, have a variety of functions in anaerobic and photosynthetic bacteria, cyanobacteria, and green algae. Their functions are similar to those of the ferredoxins, iron-sulfur proteins that are considered in Chapter 16. 

The enzyme is organized into two structural domains,201 one of which binds FAD and the other NADP+. Similar single-electron transfers through flavoproteins also occur in many other enzymes. Chorismate mutase, an important enzyme in biosynthesis of aromatic rings (Chapter 25), contains bound FMN. Its function is unclear but involves formation of a neutral flavin radical.276 277... 

The reaction is more complex than it appears. As soon as a small amount of oxidized flavin is formed, it reacts with reduced flavin to generate flavin radicals F1H (Eq. 15-27). The latter react rapidly with 02 each donating an electron to form superoxide anion radicals 02 (Eq. 15-30a) which can then combine with flavin radicals (Eq. 15-30b).284... 

Flavin adenine dinucleotide. See FAD Flavin adenine diphosphate. See FAD Flavin coenzymes 766,780 - 795 modified 788, 789 reduced 794 Flavin radicals 792 color of 794 formation constant 794 Flavocytochrome b2 782, 794, 847 Flavodoxins 793, 799, 800 Flavoprotein(s) 513, 788... 

Engelmann MD, Bobier RT, Hiatt T, Cheng IF (2003) Variability of the Fenton reaction characteristics of the EDTA, DTPAand citrate complexes of iron. BioMetals 16 519-527 Epsztejn S, Kakhlon O, Glickstein H, Breuer W, Cabantchik I (1997) Fluorescence analysis of the labile iron pool of mammalian cells. Anal Biochem 248 31-40 Faraggi M, Hemmerich P, Pecht I (1975) Oj-affinity of flavin radical species as studied by pulse radiolysis. FEBS Lett 51 47-51... 

Ulanski P, von Sonntag C (2000) Stability constants and decay of aqua-copper(lll) - a study by pulse radiolysis with conductometric detection. Eur J Inorg Chem 1211-1217 Vaish SP, Tollin G (1971) Flash photolysis of flavins. V. Oxidation and disproportionation of flavin radicals. J Bioenergetics 2 61-72... 

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