Electron-transfer oxidoreductase

Further improvements can be achieved by replacing the oxygen with a non-physiological (synthetic) electron acceptor, which is able to shuttle electrons from the flavin redox center of the enzyme to the surface of the working electrode. Glucose oxidase (and other oxidoreductase enzymes) do not directly transfer electrons to conventional electrodes because their redox center is surroimded by a thick protein layer. This insulating shell introduces a spatial separation of the electron donor-acceptor pair, and hence an intrinsic barrier to direct electron transfer, in accordance with the distance dependence of the electron transfer rate (11) ... 

Moreover, an electron transfer chain could be reconstituted in vitro that is able to oxidize aldehydes to carboxylic acids with concomitant reduction of protons and net production of dihydrogen (213, 243). The first enzyme in this chain is an aldehyde oxidoreductase (AOR), a homodimer (100 kDa) containing one Mo cofactor (MOD) and two [2Fe—2S] centers per subunit (199). The enzyme catalytic cycle can be regenerated by transferring electrons to flavodoxin, an FMN-con-taining protein of 16 kDa (and afterwards to a multiheme cytochrome and then to hydrogenase) ... 

Midpoint potential values are useful quantitites for defining the role of the various centers in the system. In some instances, these values have even been used to predict the location of the centers in the electron transfer chain, assuming that the potential increases along the chain from the electron donor to the electron acceptor. In several oxidoreductases, however, the measured potential of some centers was found to be clearly outside the range defined by the donor and the acceptor, which raised an intriguing question as to their function. This was observed, for instance, in the case of the [4Fe-4S] (Eni = -320 mV) center in E. coli fumarate reductase (249), the [3Fe-4S] + (Era = -30 mV) center in D. gigas hydrogenase (207), and the low-potential [4Fe-4S] + + (E, = 200 and -400 mV) centers in E. 

Ebenau-Jehle, M Boll, G Fuchs (2003) 2-oxoglutarate NADP oxidoreductase m Azoarcus evansii properties and function in electron transfer reactions in aromatic ring reduction. J Bacterial 185 6119-6129. 

Further research in improving the BDS activity of the biocatalysts was targeted towards the search of co-catalysts and co-factors to enhance overall desulfurization rates as well as promoters to enhance enzyme expression. This research resulted in identification of NADH and FMNH2 as co-factors essential for electron transfer and related oxidoreductase enzymes as co-catalysts as described in detail below. Additionally, other bacterial strains were also investigated as hosts and are reported below. 

NADH-coenzyme Q (CoQ) oxidoreductase, transfers electrons stepwise from NADH, through a flavoprotein (containing FMN as cofactor) to a series of iron-sulfur clusters (which will be discussed in Chapter 13) and ultimately to CoQ, a lipid-soluble quinone, which transfers its electrons to Complex III. A If, for the couple NADH/CoQ is 0.36 V, corresponding to a AG° of —69.5 kJ/mol and in the process of electron transfer, protons are exported into the intermembrane space (between the mitochondrial inner and outer membranes). 

Ferredoxins are electron-transfer proteins that can mediate between pyruvate ferredoxin oxidoreductase and hydrogenase. It appears that during the course of the evolution, different types of ferredoxin were recruited for this purpose. In Clostridia, ferredoxins of the 2[4Fe-4S] type are used (Uyeda and Rabinowitz 1971). In T. vaginalis (Chapman et al. 1986) and T. foetus (Marczak et al. 1983), [2Fe-2S] ferredoxins are used. Their axial EPR spectra at g = 1.94,2.02 (Fig. 9.2) resemble those of the ferredoxins that are involved in P450 monooxygenase systems. Similar ferredoxins, with various functions, have been isolated from... 

Ferredoxins (Fds) are widespread in the three domains of life and an abundance of sequence data and structural information are available for Fds isolated from several sources. In particular, the bacterial type Fds are small electron-transfer proteins that posses cubane xFe-yS clusters attached to the protein matrix by Fe ligation of Cys via a conserved consensus ligating sequence. The archaeal type ferredoxins are water-soluble electron acceptors for the acyl-coenzyme A forming 2-oxoacid/ferredoxin oxidoreductase, a key enzyme involved in the central archaeal metabolic pathways. Fds have been distinguished according to the number of iron and inorganic sulphur atoms, 2Fe-2S, 4Fe-4S/3Fe-4S (Fig. Ib-d) and Zn-containing Fds. 

Metal ions in the form of organometallic complexes such as the iron atom in heme can undergo one-electron transfers in oxidation-reduction reactions catalyzed by oxidoreductases with associated cytochromes. 

Pyridine nucleotide-dependent flavoenzyme catalyzed reactions are known for the external monooxygenase and the disulfide oxidoreductases However, no evidence for the direct participation of the flavin semiquinone as an intermediate in catalysis has been found in these systems. In contrast, flavin semiquinones are necessary intermediates in those pyridine nucleotide-dependent enzymes in which electron transfer from the flavin involves an obligate 1-electron acceptor such as a heme or an iron-sulfur center. Examples of such enzymes include NADPH-cytochrome P4S0 reductase, NADH-cytochrome bs reductase, ferredoxin — NADP reductase, adrenodoxin reductase as well as more complex enzymes such as the mitochondrial NADH dehydrogenase and xanthine dehydrogenase. 

Capeillere-Blandin, C., Barber, M. J., Bray, R. C. Differences in electron transfer rates among prosthetic groups between two homologous flavocytochrome b2 (L-lactate cytochrome c oxidoreductase) from different yeasts. In Flavins and flavoproteins (Massey, V., Williams, C. H. eds.) pp. 838-843, New York, Amsterdam, Oxford, Elsevier/North Holland 1982... 

FIGURE 19-9 IMADH ubiquinone oxidoreductase (Complex I). Complex I catalyzes the transfer of a hydride ion from NADH to FMN, from which two electrons pass through a series of Fe-S centers to the iron-sulfur protein N-2 in the matrix arm of the complex. Electron transfer from N-2 to ubiquinone on the membrane arm forms QH2, which diffuses into the lipid bilayer. This electron transfer also drives the expulsion from the matrix of four protons per pair of electrons. The detailed mechanism that couples electron and proton transfer in Complex I is not yet known, but probably involves a Q cycle similar to that in Complex III in which QH2 participates twice per electron pair (see Fig. 19-12). Proton flux produces an electrochemical potential across the inner mitochondrial membrane (N side negative, P side positive), which conserves some of the energy released by the electron-transfer reactions. This electrochemical potential drives ATP synthesis. 

Complex III Ubiquinone to Cytochrome c The next respiratory complex, Complex III, also called cytochrome focx complex or ubiquinone icytochrome c oxidoreductase, couples the transfer of electrons from ubiquinol (QH2) to cytochrome c with the vectorial transport of protons from the matrix to the intermembrane space. The determination of the complete structure of this huge complex (Fig. 19-11) and of Complex IV (below) by x-ray crystallography, achieved between 1995 and 1998, were landmarks in the study of mitochondrial electron transfer, providing the structural framework to integrate the many biochemical observations on the functions of the respiratory complexes. 

Another flavoprotein that makes use of both one-and two-electron transfer reactions is ferredoxin-NADP+ oxidoreductase (Eq. 15-28). Its bound FAD accepts electrons one at a time from each of the two... 

The Z scheme. [(Mn)4 = a complex of four Mn atoms bound to the reaction center of photosystem II Yz = tyrosine side chain Phe a = pheophytin a QA and Qb = two molecules of plastoquinone Cyt b/f= cytochrome hf,f complex PC = plastocyanin Chi a = chlorophyll a Q = phylloquinone (vitamin K,) Fe-Sx, Fe-SA, and Fe-SB = iron-sulfur centers in the reaction center of photosystem I FD = ferredoxin FP = flavoprotein (ferredoxin-NADP oxidoreductase).] The sequence of electron transfer through Fe-SA and Fe-SB is not yet clear. 

The molar masses of the 2-oxoacid ferredoxin oxidoreductases are 200,000-300,000 g/mol and they are composed of four subunits of the kind a2p2. It has been shown that halobacteria have only these systems of 2-oxoacid ferredoxin oxidoreductases. The two enzymes of H. halobium (pyruvate and oxoglutarate) were isolated and characterized by Kerscher and Oesterhelt (1981a). These systems proved to be thiamin diphosphate-containing iron-sulfur proteins. The relative stability of the halobacterial enzymes enabled detailed analysis of the various steps of the catalytic cycles (Kerscher and Oesterhelt, 1981b), demonstrating two distinct steps of one-electron transfer reactions. 

Many proteins are exclusively involved in intra-protein electron transfer and typically function in ordered structures such as mitochondria. Under these circumstances, the redox-active centers are generally accessible on the outer surface of the protein. In contrast, the redox reactions catalyzed by oxidoreductases involve small molecules with the reaction involving two redox couples, i.e. the substrate and the co-factor or co-substrate. Because the catalytic center of the enzyme is often located... 

Wilson s disease is a pathological accumulation of copper in tissue which is later released into the bloodstream, leading to anaemia, and final accumulation of copper in liver and brain. It is the result of a mutation in the Wilson s disease gene in chromosome 13 which ordinarily codes for a cation transporting ATPase so that copper can be incorporated into ceruloplasmin prior to excretion. Also known as ferroxi-dase, in acknowledgement of its primary function as an oxidoreductase responsible for electron transfer, this enzyme contains iron and, more importantly, six copper atoms. It accounts for the transport of 90% of copper in the plasma so any impairment in its production or efficacy has a major impact on copper homeostasis. The greatly reduced concentration of ceruloplasmin in the blood of Wilson s disease sufferers correlates with their inability to metabolize copper effectively. It leads to chronic liver disease, for which the only real cure is a liver transplant,... 

Fig. 5.2. Possible metabolic pathways in facultative anaerobic mitochondria. Shaded boxes show components of the electron-transport chain used during hypoxia, open boxes are components used during aerobiosis, and the hatched boxes (complex I and ATP-synthase) are components used under aerobic as well as anaerobic conditions. ASCT acetate succinate CoA-transferase, C cytochrome c, Cl, CIII and CIV complexes I, III and IV of the respiratory chain, CITR citrate, ECR enoyl-CoA reductase (such as present in Ascaris suum), ETF electron-transfer flavoprotein, ETF RQ OR electron-transfer flavoproteimrhodoquinone oxidoreductase, FRD fumarate reductase, FUM fumarate, MAE malate, OXAC oxaloacetate, PYR pyruvate, RQ rhodoquinone, SDH succinate dehydrogenase, SUCC succinate, Succ-CoA succinyl-CoA, TER trans-2-enoyl-CoA reductase (such as present in E. gracilis), UQ ubiquinone...

Inui H, Ono K, Miyatake K, Nakano Y, Kitaoka S (1987) Purification and characterization of pyruvate NADP+ oxidoreductase in Euglena gracilis. J Biol Chem 262 9130- 9135 Kita K, Takamiya S (2002) Electron-transfer complexes in Ascaris mitochondria. Adv Parasitol 51 95-131... 

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