Flavins protonation

Flavin coenzymes can exist in any of three different redox states. Fully oxidized flavin is converted to a semiqulnone by a one-electron transfer, as shown in Figure 18.22. At physiological pH, the semiqulnone is a neutral radical, blue in color, with a A ax of 570 nm. The semiqulnone possesses a pAl of about 8.4. When it loses a proton at higher pH values, it becomes a radical anion, displaying a red color with a A ax of 490 nm. The semiqulnone radical is particularly stable, owing to extensive delocalization of the unpaired electron across the 77-electron system of the isoalloxazine. A second one-electron transfer converts the semiqulnone to the completely reduced dihydroflavin as shown in Figure 18.22. 

Kurfuerst, M., Macheroux, P., Ghisla, S., and Hastings, J. W. (1989). Bioluminescence emission of bacterial luciferase with 1 -deaza-FMN. Evidence for the noninvolvement of N(l)-protonated flavin species as emitters. Eur. J. Biochem. 181 453 157. 

Complex II contains four peptides, the two largest form succinate dehydrogenase, the largest has covalently boiuid flavin adenine dinucleotide (FAD) which reacts with succinate, and the other has three iron-sulphur centers. Smaller subunits anchor the two larger subunits to the membrane and form the UQ binding site. Ubiquinone is the electron acceptor but complex II does not pump protons (see below). 

Alcohol dehydrogenases found in certain microorganisms utilize a pyrroloquino-line quinone (PQQ) or flavin cofactor to pass electrons released upon oxidation of alcohols to the heme electron-acceptor protein, cytochrome c. These membrane-associated alcohol dehydrogenases form part of a respiratory chain, and the energy from fuel oxidation therefore contributes to generation of a proton gradient across... 

Scheme 2 Mechanism of repair of cyclobutane pyrimidine dimers (CPD) by a CPD photolyase. 8-HDF 8-hydroxy-5-deazaflavin, ET electron transfer. FADH reduced and de-protonated flavin-coenzyme...

We next focus on the use of fixed-site cofactors and coenzymes. We note that much of this coenzyme chemistry is now linked to very local two-electron chemistry (H, CH3", CH3CO-, -NH2,0 transfer) in enzymes. Additionally, one-electron changes of coenzymes, quinones, flavins and metal ions especially in membranes are used very much in very fast intermediates of twice the one-electron switches over considerable electron transfer distances. At certain points, the chains of catalysis revert to a two-electron reaction (see Figure 5.2), and the whole complex linkage of diffusion and carriers is part of energy transduction (see also proton transfer and Williams in Further Reading). There is a variety of additional coenzymes which are fixed and which we believe came later in evolution, and there are the very important metal ion cofactors which are separately considered below. 

S0 -> Sx -> Tx transition Tt is a base of pyridine strength (pK 5), while S0 and Sj are practically non-basic (pK 0),1S9). Moreover, the site of protonation is Nl/2a for S0 andSi,butN5 forTj (Fig. 4). Hence, Hemmerich and Schmidt proposed87) that a regiospecific rearrangement of hydrogen bridges between flavin and an apoprotein environment may occur in a photo-excited flavoprotein, which would induce a unidirectional proton transfer. 

The redox and proton transfer reactions undergone by the flavin prosthetic group are summarized in Scheme 5.2. The vertical reactions are oxidations by Q regenerating P. From the standard potential values (V vs. SCE) of the four flavin redox couples that are involved in Scheme 5.2 and those of the mediators (Table 5.1), all four oxidation steps may be regarded as irreversible. The horizontal reactions are deprotonations by the bases present in the buffer. From the pA values of the various flavin acid-base couples indicated in Scheme 5.2 (over or below the horizontal arrows), reactions H2 and H4 may be regarded as irreversible and reactions HI and... 

The flavin nucleotides are typically involved in the oxidations creating double bonds from single bonds. The flavin takes up two hydrogen atoms, represented in the figure as being derived by transfer of hydride from the substrate and a proton from the medium. 

LSDl, also known as BHCllO, is the first lysine specific demethylase that was discovered. It has been assigned to group I of lysine demethylases (KDMl) [90, 91]. LSDl contains an amine oxidase domain responsible of the enzymatic activity and has been isolated as a stable component from several histone modifying complexes. The enzymatic characterization of this protein revealed that FAD (flavine adenine dinucleotide) is required as a cofactor for the removal of the methyl group. Furthermore, LSDl requires a protonated nitrogen in order to initiate demethylation so that this enzyme is only able to demethylate mono- or dimethylated substrates but not trimethylated substrates [98, 99]. 

Riboflavin (vitamin B2) is found in liver, milk, meat, green vegetables, cereals and mushrooms. It is active in the form of two coenzymes, flavin mononucleotide and flavin adenine dinucleotide. As a coenzyme for proton transfer in the respiratory chain it is indispensable for energy-release from carbohydrates, lipids and proteins. Riboflavin deficiency only occurs in combination with deficiencies of other members of the vitamin B family. The symptoms of such deficiency consist of angular stomatitis, lesions of the cornea, dermatoses and normochromic normocytic anaemia. 

Vitamin Bj (8.44, riboflavin) is a benzopteridine derivative carrying a ribityl (reduced ribose) side chain. It occurs in almost all foods, the largest amounts being found in eggs, meat, spinach, liver, yeast, and milk. Riboflavin is one of the major electron carriers as a component of flavine-adenine dinucleotide (FAD), which is involved in carbohydrate and fatty acid metabolism. A hydride ion and a proton are added to the pyrazine ring of... 

The reactions most commonly involved in flavin redox chemistry are shown in Equations 1.15-1.17. One-electron reduction of the flavin (Eq. 1.15) produces a relatively stable radical anion. Protonation of the radical anion produces an unstable neutral radical (Eq. 1.16), which will be rapidly reduced by another electron (Eq. 1.17) to give the flavohydroquinone anion. 

The structure of the modified flavocoenzymes was elucidated by chemical synthesis and comparative physical studies 126, iso) (Scheme 2, (7), (S)). The compounds possess some unusual properties some of which are collected in Table 3. The most prominent difference between (7) and (S) is the fluorescence behaviour (7) is fluorescent, (5) does not fluoresce. Moreover, at pH > 10 the fluorescence quantum yield of (7) increases by a factor of about 2, in contrast to normal flavin the fluorescence of which is quenched. By this property (7) and (5) are easily distinguished (Table 3). From the visible absorption properties of analogs of (7) and (5) it was concluded that both compounds can exist in two tautomeric forms (proton on N(l) or C(8)O, C(6)O), leading to quinoid structures. 

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