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William A. Cramer - One of the best experts on this subject based on the ideXlab platform.
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quinone dependent proton transfer pathways in the photosynthetic Cytochrome B6f complex
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Saif S Hasan, Danas Baniulis, Eiki Yamashita, William A. CramerAbstract:As much as two-thirds of the proton gradient used for transmembrane free energy storage in oxygenic photosynthesis is generated by the Cytochrome B6f complex. The proton uptake pathway from the electrochemically negative (n) aqueous phase to the n-side quinone binding site of the complex, and a probable route for proton exit to the positive phase resulting from quinol oxidation, are defined in a 2.70-A crystal structure and in structures with quinone analog inhibitors at 3.07 A (tridecyl-stigmatellin) and 3.25-A (2-nonyl-4-hydroxyquinoline N-oxide) resolution. The simplest n-side proton pathway extends from the aqueous phase via Asp20 and Arg207 (Cytochrome B6 subunit) to quinone bound axially to heme cn. On the positive side, the heme-proximal Glu78 (subunit IV), which accepts protons from plastosemiquinone, defines a route for H+ transfer to the aqueous phase. These pathways provide a structure-based description of the quinone-mediated proton transfer responsible for generation of the transmembrane electrochemical potential gradient in oxygenic photosynthesis.
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on the structural role of the aromatic residue environment of the chlorophyll a in the Cytochrome B6f complex
Biochemistry, 2008Co-Authors: Jiusheng Yan, Danas Baniulis, Naranbaatar Dashdorj, Sergei Savikhin, Eiki Yamashita, William A. CramerAbstract:Because light is not required for catalytic turnover of the Cytochrome B6f complex, the role of the single chlorophyll a in the structure and function of the complex is enigmatic. Photodamage from this pigment is minimized by its short singlet excited-state lifetime (∼200 ps), which has been attributed to quenching by nearby aromatic residues (Dashdorj et al., 2005). The crystal structure of the complex shows that the fifth ligand of the chlorophyll a contains two water molecules. On the basis of this structure, the properties of the bound chlorophyll and the complex were studied in the cyanobacterium, Synechococcus sp. PCC 7002, through site-directed mutagenesis of aromatic amino acids in the binding niche of the chlorophyll. The B6f complex was purified from three mutant strains, a double mutant Phe133Leu/Phe135Leu in subunit IV and single mutants Tyr112Phe and Trp125Leu in the Cytochrome B6 subunit. The purified B6f complex from Tyr112Phe or Phe133Leu/Phe135Leu mutants was characterized by (i) a loss o...
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chapter 20 structure function of the Cytochrome B6f complex a design that has worked for three billion years
2007Co-Authors: William A. Cramer, Jiusheng Yan, Huamin Zhang, Genji Kurisu, Naranbaatar Dashdorj, Eiki Yamashita, Hanyoup Kim, Sergei SavikhinAbstract:The 3.0–3.1 A X-ray structures of the Cytochrome B6 f complex from the thermophilic cyanobacterium Mastigocladus laminosus and from the green alga Chlamydomonas reinhardtii are very similar. Eight natural prosthetic groups, four hemes, one [2Fe-2S] cluster, one Chl, one β-carotene, and one n-side plastoquinone are embedded in the eight polypeptide subunits of the complex, four large (18–33 kDa) and four small (∼4 kDa). The complex is organized as a dimer with a molecular weight of 217 kDa in M. laminosus. Other subunits such as ferredoxin: NADP+ reductase may bind transiently and more weakly to the n-side of the complex. Major features of the structure are: (i) a large inter-monomer lipophilic “quinone exchange cavity” that exchanges plastoquinone/quinol with the quinone pool in the lipid bilayer membrane; (ii) a labyrinthine pathway of plastoquinone movement between n- and p-electron exchange sites through the 11 × 12 A portal at the roof of the cavity; (iii) three prosthetic groups with unknown function, a novel high-spin heme (cn) close to heme bn, a chlorophyll a, and a β-carotene; (iv) a proposed function of heme cn is in PS I-linked cyclic electron transport, although the presumed binding site of a “sometime” inhibitor of cyclic ET, antimycin A, is occluded by heme cn; (v) the single Chl a molecule in the monomer is characterized by a short (200 ps) fluorescence lifetime and large anisotropy of fluorescence; and (vi) transfer of energy from the Chl triplet state to the β-carotene occurs despite the 14 A separation of the pigments – it is proposed that this transfer operates through an intraprotein, interpigment O2 channel.
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intraprotein transfer of the quinone analogue inhibitor 2 5 dibromo 3 methyl 6 isopropyl p benzoquinone in the Cytochrome B6f complex
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Jiusheng Yan, Genji Kurisu, William A. CramerAbstract:Details are presented of the structural analysis of the Cytochrome B6f complex from the thermophilic cyanobacterium, Mastigocladus laminosus, in the presence of the electrochemically positive (p)-side quinone analogue inhibitor, 2,5-dibromo-3-methyl-6-isopropylbenzoquinone (DBMIB). One DBMIB binding site was found. This site is peripheral to the quinone binding space defined by the binding sites of other p-side inhibitors previously resolved in Cytochrome bc1/B6f complexes. This high-affinity site resides in a p-side interfacial niche bounded by Cytochrome f, subunit IV, and Cytochrome B6, is close (8 A) to the p-side heme b, but distant (19 A) from the [2Fe-2S] cluster. No significant electron density associated with the DBMIB was found elsewhere in the structure. However, the site at which DBMIB can inhibit light-induced redox turnover is within a few A of the [2Fe-2S] cluster, as shown by the absence of inhibition in mutants of Synechococcus sp. PCC 7002 at iron sulfur protein-Leu-111 near the cluster. The ability of a minimum amount of initially oxidized DBMIB to inhibit turnover of WT complex after a second light flash implies that there is a light-activated movement of DBMIB from the distal peripheral site to an inhibitory site proximal to the [2Fe-2S] cluster. Together with the necessary passage of quinone/quinol through the small Qp portal in the complex, it is seen that transmembrane traffic of quinone-like molecules through the core of Cytochrome bc complexes can be labyrinthine.
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characterization of the high spin heme x in the Cytochrome B6f complex of oxygenic photosynthesis
Biochemistry, 2004Co-Authors: Huijuan Zhang, Andrew Primak, John Cape, Michael K. Bowman, David Kramer, William A. CramerAbstract:X-ray structures at 3.0-3.1 A resolution of the Cytochrome B6 f complex from the cyanobacterium Mastigocladus laminosus(Kurisu, G., Zhang, H., Smith, J. L., and Cramer, W. A. (2003) Science 302, 1009-1014) and the green alga Chlamydomonas reinhardtii(Stroebel, D., Choquet, Y., Popot, J.-L., and Picot, D. (2003) Nature 426, 413-418) showed the presence of a unique heme, heme x, that is covalently linked by a single thioether bond to a Cys residue (Cys35) on the electrochemically negative (n) side of the Cytochrome B6 polypeptide. Heme x faces the intermonomer quinone exchange cavity. The only axial ligand associated with this heme is a H2 Oo r OH - that is H-bonded to the propionate of the stromal side heme bn, showing that it is pentacoordinate. The spectral properties of this heme were hardly defined at the time of the structure determination. The pyridine hemochromagen redox difference spectrum for heme x covalently bound to the Cytochrome b polypeptide isolated from SDS-PAGE displays a low-amplitude broad spectrum with a peak at 553 nm, similar to that of other hemes with a single thioether linkage. The binding of CO and a hydrophobic cyanide analogue, butyl isocyanide, to dithionite- reduced B6 f complex perturbs and significantly shifts the redox difference visible spectrum. Together with EPR spectra displaying g values of the oxidized complex of 6.7 and 7.4, heme x is defined as a ferric high-spin heme in a rhombic environment. In addition to a possible function in photosystem I-linked cyclic electron transport, the five-coordinate state implies that there is at least one more function of heme x that is related to axial binding of a physiological ligand.
Norman P A Huner - One of the best experts on this subject based on the ideXlab platform.
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Adaptation to Low Temperature in a Photoautotrophic Antarctic Psychrophile, Chlamydomonas sp. UWO 241
Photosynthesis: Structures Mechanisms and Applications, 2017Co-Authors: Beth Szyszka, Alexander G. Ivanov, Norman P A HunerAbstract:Permanent cold environments account for a large portion of the Earth. These environments are inhabited by various micro-organisms that have often adapted to unique combinations of selection pressures. Therefore, there is considerable interest in understanding survival strategies utilized by extremophiles to exist in such harsh environments. This chapter summarizes common adaptive mechanisms of psychrophilic organisms with focus on the unique photosynthetic characteristics of a unicellular green microalga, Chlamydomonas sp. UWO241. Chlamydomonas sp. UWO 241 was isolated from perennially, ice-covered Lake Bonney, Antarctica where it has adapted to constant low temperatures and high salinity. A unique characteristic of this algal strain is its inability to undergo state transitions combined with its high rates of photosystem I cyclic electron transport. Consequently, in contrast to mesophilic green algal species such as Chlamydomonas reinhardtii which undergo state transitions, Chlamydomonas sp. UWO241 does not phosphorylate LHCII polypeptides. Rather, the Antarctic psychrophile exhibits a unique, light-dependent, thylakoid polypeptide phosphorylation profile associated with a photosystem I supercomplex which also contains the Cytochrome B6/f complex. The stability of this photosystem I supercomplex in Chlamydomonas sp. UWO 241 is sensitive to its phosphorylation status as well as high salt concentrations. The role of the photosystem I supercomplex and its phosphorylation status in the regulation of photosystem I cyclic electron transport is discussed. We suggest that Chlamydomonas sp. UWO 241 should be considered a model system to study psychrophily and adaptation to low temperature in eukaryotic photoautrophs.
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the antarctic psychrophile chlamydomonas sp uwo 241 preferentially phosphorylates a photosystem i Cytochrome B6 f supercomplex
Plant Physiology, 2015Co-Authors: Beth Szyszkamroz, Alexander G. Ivanov, Paula Pittock, Gilles A Lajoie, Norman P A HunerAbstract:Chlamydomonas sp. UWO 241 (UWO 241) is a psychrophilic green alga isolated from Antarctica. A unique characteristic of this algal strain is its inability to undergo state transitions coupled with the absence of photosystem II (PSII) light-harvesting complex protein phosphorylation. We show that UWO 241 preferentially phosphorylates specific polypeptides associated with an approximately 1,000-kD pigment-protein supercomplex that contains components of both photosystem I (PSI) and the Cytochrome B6/f (Cyt B6/f) complex. Liquid chromatography nano-tandem mass spectrometry was used to identify three major phosphorylated proteins associated with this PSI-Cyt B6/f supercomplex, two 17-kD PSII subunit P-like proteins and a 70-kD ATP-dependent zinc metalloprotease, FtsH. The PSII subunit P-like protein sequence exhibited 70.6% similarity to the authentic PSII subunit P protein associated with the oxygen-evolving complex of PSII in Chlamydomonas reinhardtii. Tyrosine-146 was identified as a unique phosphorylation site on the UWO 241 PSII subunit P-like polypeptide. Assessment of PSI cyclic electron transport by in vivo P700 photooxidation and the dark relaxation kinetics of P700+ indicated that UWO 241 exhibited PSI cyclic electron transport rates that were 3 times faster and more sensitive to antimycin A than the mesophile control, Chlamydomonas raudensis SAG 49.72. The stability of the PSI-Cyt B6/f supercomplex was dependent upon the phosphorylation status of the PsbP-like protein and the zinc metalloprotease FtsH as well as the presence of high salt. We suggest that adaptation of UWO 241 to its unique low-temperature and high-salt environment favors the phosphorylation of a PSI-Cyt B6/f supercomplex to regulate PSI cyclic electron transport rather than the regulation of state transitions through the phosphorylation of PSII light-harvesting complex proteins.
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The small domain of Cytochrome f from the psychrophile Chlamydomonas raudensis UWO 241 modulates the apparent molecular mass and decreases the accumulation of Cytochrome f in the mesophile Chlamydomonas reinhardtii.
Biochemistry and Cell Biology, 2007Co-Authors: Loreta Gudynaite-savitch, Christelle Loiselay, John Simmondsj. Simmonds, Susanne E. Kohalmi, Yves Choquety. Choquet, Leonid V Savitch, Norman P A HunerAbstract:Cytochrome f from the psychrophile Chlamydomonas raudensis UWO 241 has a lower thermostability of its c- type heme and an apparent molecular mass that is 7 kDa lower than that of the model mesophilic green alga Chlamydomo- nas reinhardtii. We combined chloroplast transformation, site-directed mutagensis, and the creation of chimeric fusion constructs to assess the contribution of specific domains and (or) amino acids residues to the structure, stability, and accu- mulation of Cytochrome f, as well as its function in photosynthetic intersystem electron transport. We demonstrate that dif- ferences in the amino acid sequence of the small domain and specific charged amino acids in the large domain of Cytochrome f alter the physical properties of this protein but do not affect either the thermostability of the c-type heme, the apparent half-life of Cytochrome f in the presence of the chloroplastic protein synthesis inhibitor chloramphenicol, or the capacity for photosynthetic intersystem electron transport, measured as e - /P700. However, pulse-labeling with ( 14 C)ace- tate, combined with immunoblotting, indicated that the negative autoregulation of Cytochrome f accumulation observed in mesophilic C. reinhardtii transformed with chimeric constructs from the psychrophile was likely the result of the defective association of the chimeric forms of Cytochrome f with the other subunits of the Cytochrome B6/f complex native to the C. reinhardtii wild type. These results are discussed in terms of the unique fatty acid composition of the thylakoid membranes of C. raudensis UWO 241 adapted to cold environments.
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The Antarctic psychrophile, Chlamydomonas subcaudata, is deficient in state I-state II transitions
Planta, 2001Co-Authors: Rachael M. Morgan-kiss, Alexander G. Ivanov, Norman P A HunerAbstract:State I–State II transitions were monitored in vivo and in vitro in the Antarctic, psychrophillic, green alga, Chlamydomonas subcaudata, as changes in the low-temperature (77 K) chlorophyll fluorescence emission maxima at 722 nm (F722) relative to 699 nm (F699). As expected, the control mesophillic species, Chlamydomonas reinhardtii, was able to modulate the light energy distribution between photosystem II and photosystem I in response to exposure to four different conditions: (i) dark/anaerobic conditions, (ii) a change in Mg2+ concentration, (iii) red light, and (iv) increased incubation temperature. This was correlated with the ability to phosphorylate both of its major light-harvesting polypeptides. In contrast, exposure of C. subcaudata to the same four conditions induced minimum alterations in the 77 K fluorescence emission spectra, which was correlated with the ability to phosphorylate only one of its major light-harvesting polypeptides. Thus, C. subcaudata appears to be deficient in the ability to undergo a State I–State II transition. Functionally, this is associated with alterations in the apparent redox status of the intersystem electron transport chain and with higher rates of photosystem I cyclic electron transport in the psychrophile than in the mesophile, based on in vivo P700 measurements. Structurally, this deficiency is associated with reduced levels of Psa A/B relative to D1, the absence of specific photosystem I light-harvesting polypeptides [R.M. Morgan et al. (1998) Photosynth Res 56:303–314] and a Cytochrome B6/f complex that exhibits a form of Cytochrome f that is approximately 7 kDa smaller than that observed in C. reinhardtii. We conclude that the Antarctic psychrophile, C. subcaudata, is an example of a natural variant deficient in State I–State II transitions.
C De Vitry - One of the best experts on this subject based on the ideXlab platform.
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MUTATIONAL ANALYSIS OF THE RIESKE IRON-SULFUR PROTEIN AND THE Cytochrome bf COMPLEX QUINONE-REDUCTASE SITE
2014Co-Authors: Yexin Ouyang, Giovanni Finazzi, C De Vitry, John Whitmarsh, Richard Grebe, Lian-wang Guo, Matthew Nelson, Qingjun Wang, Claudia Facciotti, Toivo KallasAbstract:The Cytochrome bf complex catalyzes the rate-limiting quinol-oxidation step of oxygenic photosynthesis, establishes a transmembrane gradient of protons for ATP synthesis, and has been implicated in redox-mediated signaling and oxygen radical production. Four major subunits (Cytochrome B6, subunit IV, the Rieske iron-sulfur protein (ISP), and Cytochrome f), four well-known prosthetic groups (two b-hemes, the Rieske 2Fe-2S cluster, and a c-type Cytochrome), and four small subunits (PetG, PetL, PetM, PetN) comprise the bf complex. Recent X–ray crystal structures reveal an additional, unique heme designated heme x in the cyanobacterium, Mastigocladus laminonsus [1] or heme c ’ in the alga, Chlamydomonas reinhardtii [2]. The extra heme, a chlorophyll, and a carotenoid per monomer are among the unique features that distinguish Cytochrome bf from bacterial/mitochondrial Cytochrome bc complexes. Data will be presented on in vitro and in vivo mutational studies of catalysis, assembly, and roles in signaling of the Chlamydomonas Rieske ISP and on the quinone-reductase (Qi) site and oxygen radical production in the cyanobacterium Synechococcus sp. PCC 7002. ANALYSIS OF THE RIESKE ISP DISULFIDE BRIDGE AND TYROSINE 87 IN VITR
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thylakoid ftsh protease contributes to photosystem ii and Cytochrome B6f remodeling in chlamydomonas reinhardtii under stress conditions
The Plant Cell, 2014Co-Authors: Alizee Malnoe, Jacqueline Girardbascou, Francis-andré Wollman, Fei Wang, C De VitryAbstract:FtsH is the major thylakoid membrane protease found in organisms performing oxygenic photosynthesis. Here, we show that FtsH from Chlamydomonas reinhardtii forms heterooligomers comprising two subunits, FtsH1 and FtsH2. We characterized this protease using FtsH mutants that we identified through a genetic suppressor approach that restored phototrophic growth of mutants originally defective for Cytochrome B6f accumulation. We thus extended the spectrum of FtsH substrates in the thylakoid membranes beyond photosystem II, showing the susceptibility of Cytochrome B6f complexes (and proteins involved in the ci heme binding pathway to Cytochrome B6) to FtsH. We then show how FtsH is involved in the response of C. reinhardtii to macronutrient stress. Upon phosphorus starvation, photosynthesis inactivation results from an FtsH-sensitive photoinhibition process. In contrast, we identified an FtsH-dependent loss of photosystem II and Cytochrome B6f complexes in darkness upon sulfur deprivation. The D1 fragmentation pattern observed in the latter condition was similar to that observed in photoinhibitory conditions, which points to a similar degradation pathway in these two widely different environmental conditions. Our experiments thus provide extensive evidence that FtsH plays a major role in the quality control of thylakoid membrane proteins and in the response of C. reinhardtii to light and macronutrient stress.
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A novel pathway of Cytochrome c biogenesis is involved in the assembly of the Cytochrome B6f complex in arabidopsis chloroplasts.
Journal of Biological Chemistry, 2008Co-Authors: Lina Lezhneva, Richard Kuras, Geneviève Ephritikhine, C De VitryAbstract:We recently characterized a novel heme biogenesis pathway required for heme c(i)' covalent binding to Cytochrome B6 in Chlamydomonas named system IV or CCB (cofactor assembly, complex C (B6f), subunit B (PetB)). To find out whether this CCB pathway also operates in higher plants and extend the knowledge of the c-type Cytochrome biogenesis, we studied Arabidopsis insertion mutants in the orthologs of the CCB genes. The ccb1, ccb2, and ccb4 mutants show a phenotype characterized by a deficiency in the accumulation of the subunits of the Cytochrome B6f complex and lack covalent heme binding to Cytochrome B6. These mutants were functionally complemented with the corresponding wild type cDNAs. Using fluorescent protein reporters, we demonstrated that the CCB1, CCB2, CCB3, and CCB4 proteins are targeted to the chloroplast compartment of Arabidopsis. We have extended our study to the YGGT family, to which CCB3 belongs, by studying insertion mutants of two additional members of this family for which no mutants were previously characterized, and we showed that they are not functionally involved in the CCB system. Thus, we demonstrate the ubiquity of the CCB proteins in chloroplast heme c(i)' binding.
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function directed mutagenesis of the Cytochrome B6f complex in chlamydomonas reinhardtii involvement of the cd loop of Cytochrome B6 in quinol binding to the q o site
Biochemistry, 1997Co-Authors: Giovanni Finazzi, Fabrice Rappaport, Pierre Joliot, C De Vitry, Sylvie Buschlen, F A WollmanAbstract:The FUD2 mutant from the green alga Chlamydomonas reinhardtii expresses a Cytochrome B6 variant of higher apparent molecular mass [Lemaire et al. (1986) Biochim. Biophys. Acta 851, 239-248]. Here, we show that the mutation corresponds to a 36 base pair duplication in the chloroplast petB gene, which corresponds to a 12 amino acid duplication in the cd loop of Cytochrome B6. The resulting protein still binds its heme cofactors and assembles into Cytochrome B6f complexes, which accumulate in wild type amounts in exponentially growing cells of FUD2. However, these Cytochrome B6f complexes show loosened binding of the Rieske protein and are more prone to degradation in aging cells. Electron transfer through the Cytochrome B6f complexes is about 8 times slower in FUD2 than in wild type cells. This is due to a slower oxidation of plastoquinol at the Q(o) site, the folding of which is most likely altered by the duplication. By varying the redox state of the plastoquinone pool in vivo, we show that there is a dramatic decrease in the affinity of the Q(o) site for plastoquinols, which is about 100 times lower in FUD2 than in wild type cells. Our results show that the value of the binding constant of plastoquinol to the Q(o) site (2 x 10(4) M(-1)) derived in [Kramer et al. (1994) Biochim. Biophys. Acta 1184, 251-262] may be extrapolated to in vivo conditions.
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Membrane association of Cytochrome B6f subunits. The Rieske iron-sulfur protein from Chlamydomonas reinhardtii is an extrinsic protein.
Journal of Biological Chemistry, 1994Co-Authors: Cécile Breyton, C De Vitry, Jeanluc PopotAbstract:The mode of membrane attachment of five subunits from Chlamydomonas reinhardtii Cytochrome B6f complex has been studied using biochemical approaches. Antisera specific for Cytochrome f, Cytochrome B6, the Rieske iron-sulfur protein, subunit IV, and a 4-kDa subunit (product of the petG gene) were used to quantify the degree of extraction of each of these polypeptides following various treatments. In contrast to the other four subunits, the Rieske protein was extracted to extents varying between 50 and 100% following two cycles of freezing and thawing in the presence of chaotropic agents (KSCN, urea, or NaI). The Rieske protein was not extracted by 2 M NaCl and was rather resistant to alkaline treatments, being extracted by 20 mM 3-(cyclohexylamino)propanesulfonic acid buffer only at pH > 11.5. The hydrodynamic behavior of the isolated Rieske protein was examined in the absence and presence of detergent by ultracentrifugation and by molecular sieving. The extracted protein bound neither to laurylmaltoside nor to C12E8 micelles. Its sedimentation coefficient (D20,w = 9.6 x 10(-11) m2 x s-1), diffusion coefficient (s20,w = 2S), an deduced molecular mass (20.0 +/- 1.7 kDa) are those expected for the monomeric protein. We conclude that the Rieske protein is extrinsic and therefore does not cross the membrane, although its association with the rest of the complex involves primarily hydrophobic interactions, and that the other four subunits analyzed are intrinsic.
Matthias Rogner - One of the best experts on this subject based on the ideXlab platform.
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ssr2998 of synechocystis sp pcc 6803 is involved in regulation of cyanobacterial electron transport and associated with the Cytochrome B6f complex
Journal of Biological Chemistry, 2006Co-Authors: Dirk Schneider, Stephanolav Wenk, Thomas Volkmer, Gabor Bernat, Helmut Kirchhoff, Matthias RognerAbstract:To analyze the function of a protein encoded by the open reading frame ssr2998 in Synechocystis sp. PCC 6803, the corresponding gene was disrupted, and the generated mutant strain was analyzed. Loss of the 7.2-kDa protein severely reduced the growth of Synechocystis, especially under high light conditions, and appeared to impair the function of the Cytochrome B6 f complex. This resulted in slower electron donation to Cytochrome f and photosystem 1 and, concomitantly, over-reduction of the plastoquinone pool, which in turn had an impact on the photosystem 1 to photosystem 2 stoichiometry and state transition. Furthermore, a 7.2-kDa protein, encoded by the open reading frame ssr2998, was co-isolated with the Cytochrome B6 f complex from the cyanobacterium Synechocystis sp. PCC 6803. ssr2998 seems to be structurally and functionally associated with the Cytochrome B6 f complex from Synechocystis, and the protein could be involved in regulation of electron transfer processes in Synechocystis sp. PCC 6803.
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electron microscopic structural analysis of photosystem i photosystem ii and the Cytochrome B6 f complex from green plants and cyanobacteria
Journal of Bioenergetics and Biomembranes, 1994Co-Authors: Egbert J Boekema, Jan P Dekker, Arjen F Boonstra, Matthias RognerAbstract:Electron microscopy (EM) in combination with image analysis is a powerful technique to study protein structure at low- and high resolution. Since electron micrographs of biological objects are very noisy, substantial improvement of image quality can be obtained by averaging individual projections. Crystallographic and noncrystallographic averaging methods are available and have been applied to study projections of the large protein complexes embedded in photosynthetic membranes from cyanobacteria and higher plants. Results of EM on monomeric and trimeric Photosystem I complexes, on monomeric and dimeric Photosystem II complexes, and on the monomeric Cytochrome B6/f complex are discussed.
Dirk Schneider - One of the best experts on this subject based on the ideXlab platform.
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heme binding properties of heterologously expressed spinach Cytochrome B6 implications for transmembrane b type Cytochrome formation
FEBS Letters, 2007Co-Authors: Carolin Dreher, Alexander Prodohl, Mathias Weber, Dirk SchneiderAbstract:In vivo and in vitro requirements for the formation of Cytochrome b(6) were examined to analyze the mechanisms of transmembrane b-type Cytochrome formation. After heterologous expression of spinach Cytochrome b(6), formation of the holo-Cytochrome was observed within the E. coli inner membrane. The transmembrane orientation of Cytochrome b(6) appeared not to be critical for heme binding and holo-Cytochrome formation. Furthermore, in vitro reconstitution of Cytochrome b(6) was possible under oxidizing as well as under reducing conditions. Taken together these observations strongly indicate that transmembrane b-type Cytochromes can spontaneously assemble in vitro as well as in a membrane.
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ssr2998 of synechocystis sp pcc 6803 is involved in regulation of cyanobacterial electron transport and associated with the Cytochrome B6f complex
Journal of Biological Chemistry, 2006Co-Authors: Dirk Schneider, Stephanolav Wenk, Thomas Volkmer, Gabor Bernat, Helmut Kirchhoff, Matthias RognerAbstract:To analyze the function of a protein encoded by the open reading frame ssr2998 in Synechocystis sp. PCC 6803, the corresponding gene was disrupted, and the generated mutant strain was analyzed. Loss of the 7.2-kDa protein severely reduced the growth of Synechocystis, especially under high light conditions, and appeared to impair the function of the Cytochrome B6 f complex. This resulted in slower electron donation to Cytochrome f and photosystem 1 and, concomitantly, over-reduction of the plastoquinone pool, which in turn had an impact on the photosystem 1 to photosystem 2 stoichiometry and state transition. Furthermore, a 7.2-kDa protein, encoded by the open reading frame ssr2998, was co-isolated with the Cytochrome B6 f complex from the cyanobacterium Synechocystis sp. PCC 6803. ssr2998 seems to be structurally and functionally associated with the Cytochrome B6 f complex from Synechocystis, and the protein could be involved in regulation of electron transfer processes in Synechocystis sp. PCC 6803.
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functional implications of pigments bound to a cyanobacterial Cytochrome B6f complex
FEBS Journal, 2005Co-Authors: Stephanolav Wenk, Dirk Schneider, U Boronowsky, Cornelia Jager, Christof Klughammer, Frank L De Weerd, Henny Van Roon, Wim F J Vermaas, Jan P DekkerAbstract:A highly purified Cytochrome B6f complex from the cyanobacterium Synechocystis sp. PCC 6803 selectively binds one chlorophyll a and one carotenoid in analogy to the recent published structure from two other B6f complexes. The unknown function of these pigments was elucidated by spectroscopy and site-directed mutagenesis. Low-temperature redox difference spectroscopy showed red shifts in the chlorophyll and carotenoid spectra upon reduction of Cytochrome B6, which indicates coupling of these pigments with the heme groups and thereby with the electron transport. This is supported by the correlated kinetics of these redox reactions and also by the distinct orientation of the chlorophyll molecule with respect to the heme cofactors as shown by linear dichroism spectroscopy. The specific role of the carotenoid echinenone for the Cytochrome B6f complex of Synechocystis 6803 was elucidated by a mutant lacking the last step of echinenone biosynthesis. The isolated mutant complex preferentially contained a carotenoid with 0, 1 or 2 hydroxyl groups (most likely 9-cis isomers of β-carotene, a monohydroxy carotenoid and zeaxanthin, respectively) instead. This indicates a substantial role of the carotenoid – possibly for strucure and assembly – and a specificity of its binding site which is different from those in most other oxygenic photosynthetic organisms. In summary, both pigments are probably involved in the structure, but may also contribute to the dynamics of the Cytochrome B6f complex.
