Rieske Fe-S protein

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. 

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. 

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). 

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. 

Application of the impure bef complex to the hydroxyapatite column yielded a similar profile as in Fig. 2b, however with a predominant cyt.bef-peak (not shown). The four subunits of about 38, 24, 19 and 15 kDa (Table I) are likely to be cytochrome f- and cytochrome be-binding subunit, the Rieske Fe-S-protein and subunit IV, respectively, consistent with the subunit composition of other cyanobacterial cyt.bef-complexes. The presence of b-type and c-type cytochromes in a ratio of 2 1 was further confirmed by reduced-minus-oxidised difference spectra (not shown). 

CHARACTERISATION OF A FULL-LENGTH cDNA CLONE ENCODING THE PEA RIESKE Fe-S PROTEIN IMPORT AND PROCESSING BY ISOLATED CHLOROPLASTS. 

The cytochrome b-f complex of the chloroplast thylakoid membrane operates as a plastoquinol-plastocyanin oxidoreductase and is thus analogous to the mitochondrial cytochrome bc complex with which it bears a large amount of structural and functional similarity. It consists in all higher plants of four components, which are cytochrome f, cytochrome b-563 the Rieske Fe-S protein, and a 17kDa... 

The aim of the work described in this paper was to study the uptake and assembly of the Rieske Fe-S protein in pea. An isolated full-length cDNA was used to direct transcription and translation in vitro, in order to produce a precursor protein for uptake and processing experiments. 

Figure 1. Comparison of pea(Ps) and spinach (So) chloroplast Rieske Fe-S proteins...

Characterisation of a Full-Length cDNA Qone Encoding the Pea Rieske Fe-S Protein ... 

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