Rieske FeS

Efforts toward developing synthetic models for the Rieske Fe-S centers focussed initially on preparing Fe2S2 cores with non-thlolate ligands, and have centered on nitrogenous ligands since the realization of their probable occurrence in the Rieske protein. In addition to the [Fe2S2(0Ar) ] " ions (Ar = aryl)... 

Figure 9. A schematic view of the probable structure of the Rieske Fe-S center (based on the data in ref. 47.)...

Dihydroxyacid dehydratase of the branched-chain amino acid biosynthetic pathway catalyzes the dehydration and tautomerization of 2,3-dihydroxy-3-methyl-(butyrate and pentanoate) to 2-keto-3-methyl(butyrate and pentanoate). The enzyme isolated from spinach recently has been shown to have not a [4Fe-4S] cluster, but rather a spectroscopically unusual [2Fe-2S] cluster in its active site (68,69). The EPR spectrum of the reduced enzyme is similar to that seen for Rieske Fe-S proteins (71) with a g-average of 1.91. Upon addition of substrate the g-average of the EPR spectrum shifts to 1.96 (opposite the effect of substrate on aconitase), and then reverts back to a g-average of 1.90 when only the product is present The dramatic changes in the EPR spectra upon addition of substrate suggest, in analogy to aconitase, that the Fe-S cluster may be directly involved in catalysis. 

Cis-dihydroxylation of aromatic substrates is catalysed by naphthalene 1,2-dioxygenase (NDO), which comprises a Rieske Fe-S cluster and non-haem iron... 

Complex III crystallizes in two distinct conformations (not shown). In one, the Rieske Fe-S center is close to its electron acceptor, the heme of cytochrome c, but relatively distant from cytochrome b and the QH2-binding site at which the Rieske Fe-S center receives electrons. In the other, the Fe-S center has moved away from cytochrome c, and toward cytochrome b. The Rieske protein is thought to oscillate between these two conformations as it is reduced, then oxidized. 

QH2 donates one electron (via the Rieske Fe-S center) to cytochrome c, and one electron (via cytochrome b) to a molecule of Q near the n side, reducing it in two steps to QH2. This reduction also uses two protons per Q, which are taken up from the matrix. 

The simpler cytochrome bc] complexes of bacteria such as E. coli,102 Paracoccus dentrificans,116 and the photosynthetic Rhodobacter capsulatus117 all appear to function in a manner similar to that of the large mitochondrial complex. The bc] complex of Bacillus subtilis oxidizes reduced menaquinone (Fig. 15-24) rather than ubiquinol.118 In chloroplasts of green plants photochemically reduced plastoquinone is oxidized by a similar complex of cytochrome b, c-type cytochrome /, and a Rieske Fe-S protein.119 120a This cytochrome b6f complex delivers electrons to the copper protein plastocyanin (Fig. 23-18). 

Molik, S., Karnauchov, I., Weidlich, C., Herrmann, R. G., and Klosgen, R. B. (2001). The Rieske Fe/S protein of the cytochrome be,// complex in chloroplasts missing link in the evolution of protein transport pathways in chloroplasts J. Biol. Chem. 276, 42761-42766. 

This was the first complete structure of the bc complex. The structure provided information about all 11 subunits and revealed that subunit 9, the mitochondrial targeting presequence of ISP, exists between two core subunits, which are most likely a mitochondrial targeting presequence peptidase. We have solved the structures of the bc complex in two different crystal forms. Surprisingly, the conformation of the Rieske FeS protein was totally different between two crystal forms, and this provided a crucial insight of the electron bifurcation mechanism at the Qp site (see Section II,F). 

Figure 10 shows the proposed ubiquinol oxidation and electron bifurcation mechanism at Qp site. (A) In the absence of the ubiquinone, the side chain of Glu-271 is connected to the solvent in the mitochondrial intermembrane space via a water chain. (B) As a reduced ubiquinol molecule binds to the site, the side chain of Glu-271 flips to form a hydrogen bond to the bound ubiquinone. (C) Now, the ISP, which is moving around the intermediate position by thermal motion is trapped at the b" position by a hydrogen bond to the bound ubiquinone. (D,E) Coupled to deprotonation, the first electron transfer occurs. Since the Rieske FeS cluster has a much higher redox potential (ca. +300 mV) than heme bl (ca. 0 mV), the first electron is favorably transferred to ISP. This yields ubisemiquinone, (F,G). After ubisemiquinone formation, the hydrogen bond to the His-161 of ISP is destabilized. The ISP moves to the c position, where the electron is transferred from the Rieske FeS cluster to heme c. Now unstable ubisemiquinone is left in the Qp pocket. The redox potential of the deprotonated ubisemiquinone is assumed to be several hundred millivolts. Now the electron transfer to the heme bl is a downhill reaction. (H) Coupled to the second electron transfer, the second proton is transferred to Glu-271 and subsequently to the mitochondrial intermembrane space. The fully oxidized ubiquinone is released to the membrane. 

As is indicated in Table 5-3, P680, P70o> the cytochromes, plastocyanin, and ferredoxin accept or donate only one electron per molecule. These electrons interact with NADP+ and the plastoquinones, both of which transfer two electrons at a time. The two electrons that reduce plastoquinone come sequentially from the same Photosystem II these two electrons can reduce the two >-hemes in the Cyt b(f complex, or a >-heme and the Rieske Fe-S protein, before sequentially going to the /-heme. The enzyme ferre-doxin-NADP+ oxidoreductase matches the one-electron chemistry of ferredoxin to the two-electron chemistry of NADP. Both the pyridine nucleotides and the plastoquinones are considerably more numerous than are other molecules involved with photosynthetic electron flow (Table 5-3), which has important implications for the electron transfer reactions. Moreover, NADP+ is soluble in aqueous solutions and so can diffuse to the ferredoxin-NADP+ oxidoreductase, where two electrons are transferred to it to yield NADPH (besides NADP+ and NADPH, ferredoxin and plastocyanin are also soluble in aqueous solutions). 

Britt, R. D., Sauer, K., Klein, M. P., Knaff, D. B., Kriauciunas, A., Yu, C. A., Yu, L., and Malkin, R., 1991, Electron spin echo envelope modulation spectroscopy supports the suggested coordination of two histidine ligands to the Rieske Fe-S centers of the cytochrome b6f complex of spinach and the cytochrome bcl complexes of Rhodospirillum rubrum, Rhodobacter sphaeroides, and bovine heart mitochondria. Biochemistry 30 1892nl901. 

Figure 18 Schematic structure of the cytochrome bc complex from mitochondria. The struemre of the complex from purple photosynthetic bacteria is thought to be similar. The pathway of electron and proton transfer (modified Q-cycle) is overlaid on the schematic structure. Movement of the Rieske FeS protein is shown by the semitransparent yellow areas ...

The Cyt bIf complex is the only electron transport complex for which the transmembrane organization of all its subunits is established. This membrane-spanning complex that functions as an intermediate electron transport complex between PS II and PS I, and translocates protons across the membrane from the stroma to the lumen, contains 4 proteins Cyt / (33 kDa), Cyt 6-563 (23 kDa), the Rieske Fe-S protein (20 kDa) and the unnamed 17 kDa protein. 

The nuclear-eiicoded Rieske Fe-S protein is not accessible to proteolytic enzymes in thylakoids [20] or right-side-out vesicles [24], but antibody labelling shows this peptide to be exposed at both thylakoid membrane surfaces [24]. Although this thylakoid gene has not yet been sequenced, it is likely to have a structure similar to that of Neurospora Fe-S protein, which has only one membrane-spanning... 

The Cyt b-f complex contains the redox components Cyt /, Cyt 6-563 and the Rieske Fe-S protein, which in spinach have been identified as polypeptides of 33, 23 and 20 kDa respectively [95,98]. The spinach complex contains in addition a polypeptide of 17 kDa with no known redox function. The reported sizes of these polypeptides estimated by SDS-gel electrophoresis vary somewhat between different laboratories, presumably beca.use of slightly different electrophoretic procedures. The estimated size of the Cyt / polypeptide varies between different plants, even when analysed in the same electrophoresis system, although the gene sequences predict polypeptides of very similar relative molecular mass. 

As with all the other photosynthetic membrane complexes, the genes for the components of the cytochrome complex are distributed between the nuclear and chlo-roplast genomes of higher plants. The chloroplast genes for the Cyt /, Cyt 6-563 and 17 kDa polypeptides have been extensively characterized, but the nuclear gene(s) for the Rieske Fe-S protein have not yet been isolated. 

The gene(s) for the Rieske Fe-S protein has not been isolated, but its location in nuclear DNA is predicted from the synthesis of a precursor form from poly(A) RNA isolated from spinach [109]. cDNA clones containing the coding sequence of the Rieske Fe-S protein should be forthcoming shortly. 

The Rieske Fe-S protein is synthesized on cytoplasmic ribosomes to give a 27 kDa precursor form [109]. This precursor is presumably required for targetting to the chloroplasts. There is no information on the regulation of expression of the gene for the Rieske Fe-S protein, nor on the assembly of the protein into the Cyt b-f complex. 

Fig. 3.1. A, The respiratory chain. Q and c stand for ubiquinone and cytochrome c, respectively. Auxiliary enzymes that reduce ubiquinone include succinate dehydrogenase (Complex II), a-glycerophosphate dehydrogenase and the electron-transferring flavoprotein (ETF) of fatty acid oxidation. Auxiliary enzymes that reduce cytochrome c include sulphite oxidase. B, Thermodynamic view of the respiratory chain in the resting state (State 4). Approximate values are calculated according to the Nernst equation using oxidoreduction states from work by Muraoka and Slater, (NAD, Q, cytochromes c c, and a oxidation of succinate [6]), and Wilson and Erecinska (b-562 and b-566 [7]). The NAD, Q, cytochrome b-562 and oxygen/water couples are assumed to equilibrate protonically with the M phase at pH 8 [7,8]. E j (A ,/ApH) for NAD, Q, 6-562, and oxygen/water are taken as —320 mV ( — 30 mV/pH), 66 mV (- 60 mV/pH), 40 mV (- 60 mV/pH), and 800 mV (- 60 mV/pH) [7-10]. FMN and the FeS centres of Complex I (except N-2) are assumed to be in redox equilibrium with the NAD/NADH couple, FeS(N-2) with ubiquinone [11], and cytochrome c, and the Rieske FeS centre with cytochrome c [10]. The position of cytochrome a in the figure stems from its redox state [6] and its apparent effective E -, 285 mV in...

The core proteins and the Rieske FeS protein can be dissociated from Triton X-lOO-solubilised cytochrome c reductase in concentrated NaCl. The cytochrome core was isolated from the dissociated Complex III of Neurospora by gel filtration [230]. It retains most of the hydrophobic character of the parent protein and seems to correspond to the membrane-embedded part of Complex III. 

Fig. 3.10. Topography of Complex III. Complex III is a dimer in the two-dimensional crystal form studied by electron microscopy. The shape of the membrane-bound enzyme particle was resolved by image reconstruction of micrographs [230]. The location of various components was predicted by comparing crystals of Complex III with those of a subcomplex lacking the Rieske FeS protein and the core proteins [222,231]. The schematic figure is adapted from Li et al. [222].

As indicated, above, the two Z -hemes of the Cyt b f complex provide a pair of reacting sites spanning the thylakoid membrane, one near the stromal side and the other near the lumenal side of the thylakoid membrane. The plastohydroquinone is first oxidized by the Rieske FeS to a semiquinone, which is then oxidized by cytochrome/, which then releases the electron to the copper protein plastocyanin. After loss of one electron by the plastohydroquinone, the resulting semiquinone loses an electron to the two fc-hemes in series. The Z -hemes operate in the so-called Q-cycle, similar to that in the mitochondrial or bacterial cytochrome bc complex, and provide a translocation of additional protons across the membrane into the lumenal space. Discussion of the cytochrome b(,f complex and the Q-cycle will be presented in Chapter 35. 

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