Three-iron centers

The data analysis presented by Emptage et al.27) for the Av Fe-S III protein was complicated by the presence of two types of centers in the same molecule. However the novel center was also proven to be present in a ferredoxin isolated from D.gigas3S). This case represents a much simpler situation since the new type of center is the only core present. We will use this example as a prototype of proteins containing the three-iron center and its properties as representative of this new type of core. The center is referred as [3 Fe—xS] (see Table 1 and 3) due to the uncertainty on the number of labile sulfur present. We note that in aconitase (a protein which also contains this new center, as we will discuss later) equal amounts of iron and labile sulfur have been detected but the values obtained are quite low due to the presence of apoprotein68). 

Fig. 9.a Mossbauer spectrum of reduced reconstituted D. gigas ferredoxin II taken at 4.2 K in a field of 600 G applied parallel to the observed 7-radiation b Mossbauer spectra of reduced D. gigas ferredoxin I (subtraction of 25 % of a spectrum of a three-iron center run in the same conditions was made). Spectrum obtained in natural abundance Fe56. Experimental conditions as for spectrum (a). 

Huynh, B. H., et al. Evidence for a Three Iron Center in a Ferredoxin from Desulfovibrio gigas, J. of Biol. Chem. in press (1980)... 

Kent, T. A. Dreyer, J.-L. Emptage, M. H. Moura, I. Moura, J. J. G. Huynh, B. H. Xavier, A. V. LeGall, J. Beinert, H. Orme-Johnson, W. H. Munck, E. Evidence for a novel three-iron center in two ferredoxins and aconitase. In Symposium on Interaction between Iron and Proteins in Oxygen and Electron Transport, Ho, C., Ed. Elsevier North Holland, Amsterdam, 1982, pp 371-374. 

The cyclic voltammogram of complex 32 is shown in Figure 20. While the CVs of complexes 23 and 24 showed single redox processes corresponding to three concurrent one-electron reductions, the CV of complex 32 showed that two distinct redox processes were occurring. These two processes can be attributed to reduction of the inner three and outer three iron centers. The Eyx values for these two redox processes occurred at -1.20 and 1.30 y respectively. This cyclic voltammogram was obtained in DMF at 40°C with a sweep rate of 0.1 V/s. 

This impressive reaction is catalyzed by stearoyl-CoA desaturase, a 53-kD enzyme containing a nonheme iron center. NADH and oxygen (Og) are required, as are two other proteins cytochrome 65 reductase (a 43-kD flavo-protein) and cytochrome 65 (16.7 kD). All three proteins are associated with the endoplasmic reticulum membrane. Cytochrome reductase transfers a pair of electrons from NADH through FAD to cytochrome (Figure 25.14). Oxidation of reduced cytochrome be, is coupled to reduction of nonheme Fe to Fe in the desaturase. The Fe accepts a pair of electrons (one at a time in a cycle) from cytochrome b and creates a cis double bond at the 9,10-posi-tion of the stearoyl-CoA substrate. Og is the terminal electron acceptor in this fatty acyl desaturation cycle. Note that two water molecules are made, which means that four electrons are transferred overall. Two of these come through the reaction sequence from NADH, and two come from the fatty acyl substrate that is being dehydrogenated. 

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). 

Finally, the three-center term,, represents the contributions of the electrons in distant bonds to the EFG at the iron centers. Their contributions can always be expected to be small. 

Owing to the almost octahedral environment of the iron center, three out of six Fe-N stretching modes are invisible in NIS and IR spectra. Those modes that transform according to and Eg representations of the ideal octahedron do not contribute to the msd of the iron nucleus or to the variation of the electric dipole moment. Only the remaining three modes that transform according to Tiu representations can be observed in NIS- and IR spectra. These three modes, as obtained... 

Under conditions of copper deficiency, some methanotrophs can express a cytosolic, soluble form of MMO (sMMO) (20-23), the properties of which form the focus of the present review. The sMMO system comprises three separate protein components which have all been purified to homogeneity (24,25). The hydroxylase component, a 251 kD protein, contains two copies each of three subunits in an a 82y2 configuration. The a subunit of the hydroxylase houses the dinuclear iron center (26) responsible for dioxygen activation and for substrate hydroxylation (27). The 38.6 kD reductase contains flavin adenine dinucleotide (FAD) and Fe2S2 cofactors (28), which enable it to relay electrons from reduced nicotinamide adenine dinucleotide (NADH) to the diiron center in the... 

FIGURE 13.2 An EPR-monitored redox titration of an Fe-O-Fe cluster with three stable oxidation states. The dinuclear iron center (= +210 mV and = +50 mV) in Pyrococcus furio-sus ferritin was titrated in the presence of a mediator mix. The fit is based on Equation 13.14. (Data from Tatur and Hagen 2005.)... 

Another complex involving the formation of a cyclopentadienyl unit is obtained from the interaction of ethyl or propyl acetylene with Fe3(CO)i2 117). The products contain the complexes Fe3(CO)7(HC2R)4 (R = Et, re-Pr), and the crystal structure of the ethyl derivative indicates the presence of the substituted 1,2,3-triethylcyclopentadienyl group bonded to one iron center with an ethylallyl group cr- and n-bonded to the three metal centers. The formation of adducts of this type must involve the fission of the C=C bond of the acetylene. 

Three other proteins with similar domain structure as that of FprA were reported in other bacteria (WasserfaUen et al. 1995 Gomes et al. 1997, 2000). The recombinant CthFprA and CthHrb, overexpressed in E. coli, were purified and characterized. Both FprA and Hrb were found to be present as dimers. Metal/cofactor analysis of the purified proteins revealed the presence of 2 mol each of iron and flavin (FMN) per mole dimer of Hrb and 4 mol of iron and 2 mol FMN per mole dimer of FprA. The EPR spectra of the purified proteins indicated that iron is present in a di-iron center in FprA and as a Fe(Cys)4 cluster in Hrb. 

Three oxidations states are potentially available in a binuclear iron center. Enzymes with octahedral fi-o o bridged iron clusters can be isolated in each of the three states the diferric and diferrous states appear to be the functional terminal oxidation states for most of the enzymes, while the mixed valence state may be an important intermediate or transition state for some reactions (Que and True, 1991). In these enzymes the cluster participates primarily as a two-electron partner in the redox of substrates, perhaps using sequential one-electron steps. Without additional coupled redox steps the enzyme is in a new oxidation state after one turnover. In contrast only the diferric and mixed valence oxidation states have been found for 2Fe 2S clusters. The diferrous state may not be obtainable because of the high negative charge on [2Fe 2S(4RS)] versus -1 or 0 net charge for the diferrous octahedral (i.e., non-Fe S) clusters. The 2Fe 2S proteins either are one-electron donor/acceptors or serve as transient electron transfer intermediates. 

A representative sampling of non-heme iron proteins is presented in Fig. 3. Evident from this atlas is the diversity of structural folds exhibited by non-heme iron proteins it may be safely concluded that there is no unique structural motif associated with non-heme iron proteins in general, or even for specific types of non-heme iron centers. Protein folds may be generally classified into several categories (i.e., all a, parallel a/)3, or antiparallel /8) on the basis of the types and interactions of secondary structures (a helix and sheet) present (Richardson, 1981). Non-heme iron proteins are found in all three classes (all a myohemerythrin, ribonucleotide reductase, and photosynthetic reaction center parallel a/)8 iron superoxide dismutase, lactoferrin, and aconitase antiparallel )3 protocatechuate dioxygenase, rubredoxins, and ferredoxins). This structural diversity is another reflection of the wide variety of functional roles exhibited by non-heme iron centers. 

The protein is composed of three independent components component A, the hydroxylase component B, a mediating protein and component C, the reductase (Green and Dalton, 1985 Fox et al., 1989). These proteins have been isolated in high purity and activity. The hydroxylase contains iron centers which have Mdssbauer parameters simi-... 

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