Membrane Association of the Rieske Iron-Sulfur Protein

Szczepaniak, Andrzej; Frank, Karin; Rybka, Jacek

doi:10.1515/znc-1995-7-811

Cited by 6 publications

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“…This experiment does not, however, distinguish between two general modes of hydrophobic anchoring through the NH 2 -terminal segment: (i) a trans-membrane ␣-helix (37) or (ii) an extrinsic protein that is released from the membrane by chaotropic agents (38) or bound within the membrane surface in the intermediate dielectric constant interfacial layer (1). The latter mode of hydrophobic association, in contrast to a trans-membrane ␣-helix, would be more readily reconciled with the inability to cleave the Rieske protein with protease probes added to the stromal side of thylakoid membranes (39). 6 f complex) and Rieske fragment (68 M, 1 mg/ml) were poised potentiometrically to oxidation-reduction potentials at pH 6.9 of ϩ217 and ϩ205 mV, respectively.…”

Section: Discussionmentioning

confidence: 99%

Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Zhang

Carrell

Huang

et al. 1996

Journal of Biological Chemistry

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A soluble, 139-residue COOH-terminal polypeptide fragment of the Rieske iron-sulfur protein of the cytochrome b 6 f complex from spinach chloroplasts was obtained by limited proteolysis of the complex and a twostep chromatography purification protocol. The purified Rieske iron-sulfur protein fragment was characterized by: (i) a single NH 2 -terminal sequence, NH 2 -Phe-Val-Pro-Pro-Gly-Gly, starting with residue 41 of the intact Rieske protein; (ii) a single molecular weight species determined by mass spectrometry with a molecular weight of 14,620 ؎ 2 without the [2Fe-2S] cluster; (iii) an optical absorbance spectrum with redox-and pHdependent maxima and minima; and (iv) a reduced-oxidized optical difference spectrum characterized by ⌬⑀ mM ‫؍‬ 3.8 mM ؊1 cm ؊1 for ⌬A at 394 versus 409 nm, which was used to determine the midpoint oxidation-reduction potential, which is ؉359 ؎ 7 mV at 25°C from pH 5.5-6.5, and ؉319 ؎ 2 mV at pH 7, with an apparent pK ox ‫؍‬ 6. The cytochrome b 6 f complex is one of three integral membrane protein complexes in the oxygenic photosynthetic membrane that participate in electron transport, H ϩ translocation, and generation of the trans-membrane proton electrochemical potential. It occupies a central position with respect to the other two integral protein systems, the photosystem II and I reaction center complexes. The Rieske high potential [2Fe-2S] iron-sulfur protein is one of four "large" polypeptides (M r ϭ 17,000 -32,000) observed in denaturing gels of the cytochrome b 6 f (plastoquinol-plastocyanin oxidoreductase) complex and one of three that contain a redox prosthetic group. In order of descending size, these four polypeptides are: cytochrome f, cytochrome b 6 , the nuclear-encoded Rieske [2Fe-2S] protein, and subunit IV, with molecular weights in the complex of spinach chloroplasts of 32,038, 24,166, 19,116, and 17,445, The first crystal structure for an active domain of the cyt b 6 f or bc 1 complexes was the 252-residue soluble lumen side fragment of the 285 residue cyt f, whose structure has been determined to a resolution of 1.96 Å (4, 5). The cytochrome was found to consist of two domains made mostly of ␤ sheet in an elongate (approximately 75 ϫ 35 ϫ 25 Å) geometry. The covalently bound heme lies within the larger domain of the elongate structure near the interface between the large and small domains, its heme iron atom 45 Å from the Arg 250 near the COOH terminus that is connected to the single trans-membrane ␣-helix. The elongate structure of cytochrome f raises questions that are defined in a structural context about the pathway of electron transfer from the membrane-bound hydrogen donor, plastoquinol, to the Rieske protein, and from the Rieske protein to cytochrome f, whose heme appears to be far from the membrane surface. In addition, the existence of a buried ordered linear H 2 O chain, a seeming proton wire with virtually all H-bond donors and acceptors conserved in evolution, which extends 11 Å from the N␦1 nitrogen of His 25 toward Lys 66 of the prominent basic su...

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Section: Discussionmentioning

confidence: 99%

Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Zhang

Carrell

Huang

et al. 1996

Journal of Biological Chemistry

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“…In contrast, the topology of the Rieske protein is a matter of considerable dispute (e.g., [14, 15]). Close to its NH 2 ‐terminus, the Rieske protein houses a stretch of 25 hydrophobic residues which might either serve to attach the protein from the lumenal side to the membrane [15, 16]or to anchor it within the membrane with one or two transmembrane spans [17, 18]. The bulk of the protein including the COOH‐terminus is located in the thylakoid lumen.…”

Section: Introductionmentioning

confidence: 99%

Transmembrane topology of the Rieske Fe/S protein of the cytochrome b₆/f complex from spinach chloroplasts

1997

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The topology of the Rieske protein of the cytochrome b 6 lf complex in thylakoids from spinach chloroplasts was examined by protease protection experiments as well as polypeptide extraction assays using solutions of chaotropic salts or alkaline pH. While neither thermolysin nor trypsin cleave any of the Rieske protein when added to the stromal side of the thylakoid membrane, proteinase K is capable of removing approximately four residues from its NH 2 -terminus. The protein is resistant to membrane extraction by 0.1 M Na 2 C0 3 or 2 M NaBr but is quantitatively released by 0.1 M NaOH. Treatment of thylakoids with 2 M NaSCN leads to extraction of variable amounts of the protein, depending on the presence or absence of sucrose in the medium which apparently stabilizes the cytochrome complex. From these results we conclude that the Rieske protein is an integral component of the cytochrome complex which spans the thylakoid membrane with a single hydrophobic segment and is anchored predominantly by electrostatic interactions.

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“…These proteins are held in the membrane by hydrophobic domains near the ends of the polypeptides; cytochrome f contains a single membrane‐spanning hydrophobic region near the C‐terminus [5], whereas the Rieske protein contains a hydrophobic region near the N‐terminus [6,7]. The way in which the Rieske protein is associated with the thylakoid membrane has been the subject of considerable controversy; it has been suggested to be an integral membrane protein with one or possibly two transmembrane spans [7,8], or to be a peripheral membrane protein associated with the lumenal face of the membrane via the hydrophobic region [9,10]. However, the recently determined structures of the mitochondrial cytochrome bc 1 complex [11–14], which has many features in common with the cytochrome bf complex, together with protease accessibility studies on chloroplast thylakoid membranes [15], suggest that the chloroplast Rieske protein contains a single transmembrane span.…”

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confidence: 99%

Assembly of the Rieske iron–sulphur protein into the cytochrome bf complex in thylakoid membranes of isolated pea chloroplasts

Kapazoglou

Mould

Gray

2000

European Journal of Biochemistry

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The assembly of the Rieske iron±sulphur protein into the cytochrome bf complex was examined following import of 35 S-labeled precursor protein by isolated pea chloroplasts. Rieske protein assembled into the cytochrome bf complex was resolved from unassembled Rieske protein and from other membrane complexes by nondenaturing gel electrophoresis of dodecyl maltoside-solubilized thylakoid membranes. Four mutant forms of the Rieske protein were able to assemble into the cytochrome bf complex in isolated chloroplasts. These were a triple substitution mutant, C107S/H109R/C112S, replacing conserved residues involved in the ligation of the [2Fe-2S] centre; the mutant D45±52 which removed a glycine-rich region predicted to form a flexible hinge between the hydrophobic membrane-associated region and the hydrophilic lumenal domain; and mutants D168±173 and D177±179 which removed two C-terminal regions, which are highly conserved in chloroplast and cyanobacterial Rieske proteins. This indicates that the [2Fe±2S] cluster, the glycine-rich region and the C-terminal region are not essential for stable assembly of the Rieske protein into the cytochrome bf complex in isolated chloroplasts.

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Membrane Association of the Rieske Iron-Sulfur Protein

Cited by 6 publications

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Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Transmembrane topology of the Rieske Fe/S protein of the cytochrome b₆/f complex from spinach chloroplasts

Assembly of the Rieske iron–sulphur protein into the cytochrome bf complex in thylakoid membranes of isolated pea chloroplasts

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Membrane Association of the Rieske Iron-Sulfur Protein

Cited by 6 publications

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Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Characterization and Crystallization of the Lumen Side Domain of the Chloroplast Rieske Iron-Sulfur Protein

Transmembrane topology of the Rieske Fe/S protein of the cytochrome b6/f complex from spinach chloroplasts

Assembly of the Rieske iron–sulphur protein into the cytochrome bf complex in thylakoid membranes of isolated pea chloroplasts

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Transmembrane topology of the Rieske Fe/S protein of the cytochrome b₆/f complex from spinach chloroplasts