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Jurgen Soll - One of the best experts on this subject based on the ideXlab platform.
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fzl is primarily localized to the inner Chloroplast Membrane however influences thylakoid maintenance
Plant Molecular Biology, 2018Co-Authors: Manali Patil, Jurgen Soll, Stephanie Seifert, Franka Seiler, Serena SchwenkertAbstract:FZL is primarily localized to the Chloroplast inner envelope and not to the thylakoids, but nevertheless affects the maintenance of thylakoid Membranes and photosynthetic protein complexes. The fuzzy-onion-like protein (FZL) is a Membrane-bound dynamin-like GTPase located in the Chloroplast. We have investigated the Chloroplast sub-localization of the endogenous FZL protein and found it to be primarily localized to the inner envelope. Moreover, we observed that mature leaves of fzl mutants start to turn pale, especially in the midvein area of the leaves, 11 days after germination. We therefore assessed their photosynthetic performance as well as the accumulation of thylakoid Membrane proteins and complexes after the initial appearance of the phenotype. Interestingly, we could observe a significant decrease in amounts of the cytochrome b (6) f complex in 20-day-old mutants, which was also reflected in an impaired electron transport rate as well as a more oxidized P700 redox state. Analysis of differences in transcriptome datasets obtained before and after onset of the phenotype, revealed large-scale changes in gene expression after the phenotype became visible. In summary, we propose that FZL, despite its localization in the inner Chloroplast envelope has an important role in thylakoid maintenance in mature and aging leaves.
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high resolution nmr reveals secondary structure and folding of amino acid transporter from outer Chloroplast Membrane
PLOS ONE, 2013Co-Authors: James Zook, Jurgen Soll, Neil E Jacobsen, Brian R Cherry, Michael F Brown, Trivikram R Molugu, Petra FrommeAbstract:Solving high-resolution structures for Membrane proteins continues to be a daunting challenge in the structural biology community. In this study we report our high-resolution NMR results for a transMembrane protein, outer envelope protein of molar mass 16 kDa (OEP16), an amino acid transporter from the outer Membrane of Chloroplasts. Three-dimensional, high-resolution NMR experiments on the 13C, 15N, 2H-triply-labeled protein were used to assign protein backbone resonances and to obtain secondary structure information. The results yield over 95% assignment of N, HN, CO, Cα, and Cβ chemical shifts, which is essential for obtaining a high resolution structure from NMR data. Chemical shift analysis from the assignment data reveals experimental evidence for the first time on the location of the secondary structure elements on a per residue basis. In addition T1Z and T2 relaxation experiments were performed in order to better understand the protein dynamics. Arginine titration experiments yield an insight into the amino acid residues responsible for protein transporter function. The results provide the necessary basis for high-resolution structural determination of this important plant Membrane protein.
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molecular properties of oep21 an atp regulated anion selective solute channel from the outer Chloroplast Membrane
Journal of Biological Chemistry, 2006Co-Authors: Roland Hemmler, Jurgen Soll, Enrico Schleiff, Thomas Becker, Bettina Bolter, Tanja Stahl, Tom A Gotze, Simona Braams, Richard WagnerAbstract:Abstract The flux of phosphorylated carbohydrates, the major export products of Chloroplasts, is regulated at the level of the inner and presumably also at the level of the outer Membrane. This is achieved through modulation of the outer Membrane Oep21 channel currents and tuning of its ion selectivity. Refined analysis of the Oep21 channel properties by biochemical and electrophysiological methods revealed a channel formed by eight β-strands with a wider pore vestibule of dvest ∼2.4 nm at the interMembrane site and a narrower filter pore of drestr ∼1 nm. The Oep21 pore contains two high affinity sites for ATP, one located at a relative transMembrane electrical distance δ = 0.56 and the second close to the vestibule at the interMembrane site. The ATP-dependent current block and reduction in anion selectivity of the Oep21 channel is relieved by the competitive binding of phosphorylated metabolic intermediates like 3-phosphoglycerate and glycerinaldehyde 3-phosphate. Deletion of a C-terminal putative FX4K binding motif in Oep21 decreased the capability of the channel to tune its ion selectivity by about 50%, whereas current block remained unchanged.
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Chloroplast Membrane transport interplay of prokaryotic and eukaryotic traits
Gene, 2005Co-Authors: Ute C Vothknecht, Jurgen SollAbstract:Abstract Chloroplasts are specific plant organelles of prokaryotic origin. They are separated from the surrounding cell by a double Membrane, which represents an effective barrier for the transport of metabolites and proteins. Specific transporters in the inner envelope Membrane have been described, which facilitate the exchange of metabolites. In contrast, the outer envelope has been viewed for a long time as a molecular sieve that offers a mere size constriction to the passage of molecules. This view has been challenged lately, and a number of specific and regulated pore proteins of the outer envelope (OEPs) have been identified. These pores seem to have originated by adaptation of outer Membrane proteins of the cyanobacterial ancestor of the Chloroplast. In a similar fashion, the transport of proteins across the two envelope Membranes is achieved by two hetero-oligomeric protein complexes called Toc (translocon in the outer envelope of Chloroplasts) and Tic (translocon in the inner envelope of Chloroplasts). The phylogenetic provenance of the translocon components is less clear, but at least the channel protein of the Toc translocon is of cyanobacterial origin. Characteristic of cyanobacteria and Chloroplasts is furthermore a specialized internal Membrane system, the thylakoids, on which the components of the photosynthetic machinery are located. Despite the importance of this Membrane, very little is known about its phylogenetic origin or the manner of its synthesis. Vipp1 appears to be a ubiquitous component of thylakoid formation, while in Chloroplasts of land plants, additionally a vesicle transport system of eukaryotic origin might be involved in this process.
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the pea Chloroplast Membrane associated protein iep96 is a subunit of acetyl coa carboxylase
Plant Journal, 1996Co-Authors: Basil S Shorrosh, Jurgen Soll, Linda J Savage, John B OhlroggeAbstract:: Two forms of acetyl-CoA carboxylase (ACCase) have been characterized in pea (Pisum sativum L.) leaves; a heteromeric Chloroplast enzyme and a homomeric, presumably cytosolic enzyme. The biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and beta-carboxyltransferase (CT) subunits of the plastidial-ACCase have recently been characterized and cloned. To further characterize the carboxyltransferase, an improved assay for CT was developed and used to follow its partial purification. CT activity co-purifies with ACCase activity during gel permeation chromatography. However, upon anion-exchange chromatography or native PAGE, CT separates from the BC and BCCP subunits of plastidial-ACCase and ACCase activity is lost. In addition, it is demonstrated that a previously sequenced pea Chloroplast cDNA of unknown function (IEP96) with a predicted molecular weight of 91 kDa encodes the alpha-CT subunit of the MS-ACCase. Antibodies raised against the first 404 amino acids of IEP96 protein detected a polypeptide with molecular weight of 91 kDa that co-eluted during gel permeation chromatography with plastidial CT and ACCase activities. These antibodies also immunoprecipitated the activities of both ACCase and CT with the concomitant precipitation of the beta-CT subunit. Furthermore, antibodies against beta-CT immunoprecipitated the IEP96 protein. Two-dimensional PAGE and DEAE purification of ACCase protein demonstrated that the beta-CT forms a tight association with the IEP96 protein. Pea leaf was fractionated into soluble and Membrane fractions and the alpha-CT subunit was primarily associated with the Membrane fraction. Together, these data demonstrate that IEP96 is the alpha-CT subunit of pea Chloroplast ACCase.
Felix Kessler - One of the best experts on this subject based on the ideXlab platform.
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the novel Chloroplast outer Membrane kinase koc1 is a required component of the plastid protein import machinery
Journal of Biological Chemistry, 2017Co-Authors: Monica Zufferey, Cyrille Montandon, Veronique Douet, Emilie Demarsy, Birgit Agne, Sacha Baginsky, Felix KesslerAbstract:Abstract The biogenesis and maintenance of cell organelles such as mitochondria and Chloroplasts require the import of many proteins from the cytosol, a process that is controlled by phosphorylation. In the case of Chloroplasts, the import of hundreds of different proteins depends on translocons at the outer and inner Chloroplast Membrane (TOC and TIC, respectively) complexes. The essential protein TOC159 functions thereby as an import receptor. It has an N-terminal acidic (A-) domain that extends into the cytosol, controls receptor specificity, and is highly phosphorylated in vivo. However, kinases that phosphorylate the TOC159 A-domain to enable protein import have remained elusive. Here, using co-purification with TOC159 from Arabidopsis, we discovered a novel component of the Chloroplast import machinery, the regulatory kinase at the outer Chloroplast Membrane 1 (KOC1). We found that KOC1 is an integral Membrane protein facing the cytosol and stably associates with TOC. Moreover, KOC1 phosphorylated the A-domain of TOC159 in vitro, and in mutant koc1 Chloroplasts, preprotein import efficiency was diminished. koc1 Arabidopsis seedlings had reduced survival rates after transfer from the dark to the light in which protein import into plastids is required to rapidly complete Chloroplast biogenesis. In summary, our data indicate that KOC1 is a functional component of the TOC machinery that phosphorylates import receptors, supports preprotein import, and contributes to efficient Chloroplast biogenesis.
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tocopherol cyclase vte1 localization and vitamin e accumulation in Chloroplast plastoglobule lipoprotein particles
Journal of Biological Chemistry, 2006Co-Authors: Pierrealexandre Vidi, Felix Kessler, Peter Dormann, Sacha Baginsky, Marion Kanwischer, Jotham R Austin, Gabor Csucs, Claire BrehelinAbstract:Abstract Chloroplasts contain lipoprotein particles termed plastoglobules. Plastoglobules are generally believed to have little function beyond lipid storage. Here we report on the identification of plastoglobule proteins using mass spectrometry methods in Arabidopsis thaliana. We demonstrate specific plastoglobule association of members of the plastid lipid-associated proteins/fibrillin family as well as known metabolic enzymes, including the tocopherol cyclase (VTE1), a key enzyme of tocopherol (vitamin E) synthesis. Moreover, comparative analysis of Chloroplast Membrane fractions shows that plastoglobules are a site of vitamin E accumulation in Chloroplasts. Thus, in addition to their lipid storage function, we propose that plastoglobules are metabolically active, taking part in tocopherol synthesis and likely other pathways.
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essential role of the g domain in targeting of the protein import receptor attoc159 to the Chloroplast outer Membrane
Journal of Cell Biology, 2002Co-Authors: Jorg Bauer, Danny J Schnell, Andreas Hiltbrunner, Mayte Alvarezhuerta, Pierrealexandre Vidi, Petra Weibel, Matthew D Smith, Felix KesslerAbstract:Two homologous GTP-binding proteins, atToc33 and atToc159, control access of cytosolic precursor proteins to the Chloroplast. atToc33 is a constitutive outer Chloroplast Membrane protein, whereas the precursor receptor atToc159 also exists in a soluble, cytosolic form. This suggests that atToc159 may be able to switch between a soluble and an integral Membrane form. By transient expression of GFP fusion proteins, mutant analysis, and biochemical experimentation, we demonstrate that the GTP-binding domain regulates the targeting of cytosolic atToc159 to the Chloroplast and mediates the switch between cytosolic and integral Membrane forms. Mutant atToc159, unable to bind GTP, does not reinstate a green phenotype in an albino mutant (ppi2) lacking endogenous atToc159, remaining trapped in the cytosol. Thus, the function of atToc159 in Chloroplast biogenesis is dependent on an intrinsic GTP-regulated switch that controls localization of the receptor to the Chloroplast envelope.
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targeting of an abundant cytosolic form of the protein import receptor at toc159 to the outer Chloroplast Membrane
Journal of Cell Biology, 2001Co-Authors: Andreas Hiltbrunner, Jorg Bauer, Pierrealexandre Vidi, Petra Weibel, Sibylle Infanger, Morten Hohwy, Felix KesslerAbstract:Chloroplast biogenesis requires the large-scale import of cytosolically synthesized precursor proteins. A trimeric translocon (Toc complex) containing two homologous GTP-binding proteins (atToc33 and atToc159) and a channel protein (atToc75) facilitates protein translocation across the outer envelope Membrane. The mechanisms governing function and assembly of the Toc complex are not yet understood. This study demonstrates that atToc159 and its pea orthologue exist in an abundant, previously unrecognized soluble form, and partition between cytosol-containing soluble fractions and the Chloroplast outer Membrane. We show that soluble atToc159 binds directly to the cytosolic domain of atToc33 in a homotypic interaction, contributing to the integration of atToc159 into the Chloroplast outer Membrane. The data suggest that the function of the Toc complex involves switching of atToc159 between a soluble and an integral Membrane form.
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protein translocon at the arabidopsis outer Chloroplast Membrane
Biochemistry and Cell Biology, 2001Co-Authors: Andreas Hiltbrunner, Jorg Bauer, Mayte Alvarezhuerta, Felix KesslerAbstract:Chloroplasts are organelles essential for the photoautotrophic growth of plants. Their biogenesis from undifferentiated proplastids is triggered by light and requires the import of hundreds of diff...
Bernhard Grimm - One of the best experts on this subject based on the ideXlab platform.
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Chloroplast Membrane photostability in chlp transgenic tobacco plants deficient in tocopherols
Plant Physiology, 2003Co-Authors: Michel Havaux, Cornelius Lutz, Bernhard GrimmAbstract:The phototolerance of three chlP transgenic tobacco ( Nicotiana tabacum ) lines, affected in geranylgeranyl reductase and, hence, deficient in tocopherols (vitamin E), was estimated by in vivo luminescence and fluorescence measurements and was compared with that of the wild type (WT). Exposure of leaf discs to high light (1 mmol photon m −2 s −1 ) and low temperature (10°C) led to a rapid inhibition of photosystem II (PSII) photochemistry that showed little dependence on the tocopherol level. PSII photo-inhibition was followed by lipid peroxidation with a time delay of about 4 h, and this phenomenon was exacerbated in the tocopherol-deficient leaves. A linear correlation was observed in these short-term experiments between resistance to photooxidation and tocopherol content. When whole plants were exposed to the same treatment, PSII was severely photo-inhibited in mature leaves of all genotypes. Lipid peroxidation was also observed in all plants, but it occurred much more rapidly in tocopherol-deficient transgenic plants relative to WT plants. The time at which extensive lipid peroxidation occurred was correlated with the tocopherol content of the leaves. The present results show that tocopherols protect thylakoid Membranes against photodestruction through lipid peroxidation. However, tocopherol deficiency was compensated in young, developing leaves that were able to photo-acclimate in the long term and did not suffer from photooxidative damage. Soluble antioxidants (glutathione and ascorbate) did not accumulate in photo-acclimated chlP transgenic leaves relative to WT leaves. In contrast, a selective accumulation of xanthophyll cycle pigments was observed in young transgenic leaves, and this could represent a compensatory mechanism for tocopherol deficiency.
Michel Havaux - One of the best experts on this subject based on the ideXlab platform.
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Chloroplast Membrane photostability in chlp transgenic tobacco plants deficient in tocopherols
Plant Physiology, 2003Co-Authors: Michel Havaux, Cornelius Lutz, Bernhard GrimmAbstract:The phototolerance of three chlP transgenic tobacco ( Nicotiana tabacum ) lines, affected in geranylgeranyl reductase and, hence, deficient in tocopherols (vitamin E), was estimated by in vivo luminescence and fluorescence measurements and was compared with that of the wild type (WT). Exposure of leaf discs to high light (1 mmol photon m −2 s −1 ) and low temperature (10°C) led to a rapid inhibition of photosystem II (PSII) photochemistry that showed little dependence on the tocopherol level. PSII photo-inhibition was followed by lipid peroxidation with a time delay of about 4 h, and this phenomenon was exacerbated in the tocopherol-deficient leaves. A linear correlation was observed in these short-term experiments between resistance to photooxidation and tocopherol content. When whole plants were exposed to the same treatment, PSII was severely photo-inhibited in mature leaves of all genotypes. Lipid peroxidation was also observed in all plants, but it occurred much more rapidly in tocopherol-deficient transgenic plants relative to WT plants. The time at which extensive lipid peroxidation occurred was correlated with the tocopherol content of the leaves. The present results show that tocopherols protect thylakoid Membranes against photodestruction through lipid peroxidation. However, tocopherol deficiency was compensated in young, developing leaves that were able to photo-acclimate in the long term and did not suffer from photooxidative damage. Soluble antioxidants (glutathione and ascorbate) did not accumulate in photo-acclimated chlP transgenic leaves relative to WT leaves. In contrast, a selective accumulation of xanthophyll cycle pigments was observed in young transgenic leaves, and this could represent a compensatory mechanism for tocopherol deficiency.
Petra Fromme - One of the best experts on this subject based on the ideXlab platform.
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high resolution nmr reveals secondary structure and folding of amino acid transporter from outer Chloroplast Membrane
PLOS ONE, 2013Co-Authors: James Zook, Jurgen Soll, Neil E Jacobsen, Brian R Cherry, Michael F Brown, Trivikram R Molugu, Petra FrommeAbstract:Solving high-resolution structures for Membrane proteins continues to be a daunting challenge in the structural biology community. In this study we report our high-resolution NMR results for a transMembrane protein, outer envelope protein of molar mass 16 kDa (OEP16), an amino acid transporter from the outer Membrane of Chloroplasts. Three-dimensional, high-resolution NMR experiments on the 13C, 15N, 2H-triply-labeled protein were used to assign protein backbone resonances and to obtain secondary structure information. The results yield over 95% assignment of N, HN, CO, Cα, and Cβ chemical shifts, which is essential for obtaining a high resolution structure from NMR data. Chemical shift analysis from the assignment data reveals experimental evidence for the first time on the location of the secondary structure elements on a per residue basis. In addition T1Z and T2 relaxation experiments were performed in order to better understand the protein dynamics. Arginine titration experiments yield an insight into the amino acid residues responsible for protein transporter function. The results provide the necessary basis for high-resolution structural determination of this important plant Membrane protein.
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secondary structure and folding of amino acid transporter from outer Chloroplast Membrane
Biophysical Journal, 2013Co-Authors: James Zook, Trivikram R Moulgu, Neil E Jacobsen, Brian R Cherry, Michael F Brown, Petra FrommeAbstract:Solving high-resolution structures for Membrane proteins continues to be a daunting challenge in the structural biology community. OEP16 is a transMembrane amino acid selective transporter with a molecular mass of 16 kDa [1]. Here we report high-resolution NMR results for OEP16 solubilized in SDS detergent micelles. Three-dimensional NMR experiments were performed on U-15N, 13C- labeled and 80% perdeuterated protein to assign protein backbone resonances and obtain secondary structure information. To understand the protein dynamics and function 15N relaxation, amino acid titration, and 2H exchange experiments were performed. The chemical shift data for 15N, 1HN, 13CO, 13Cα and 13Cβ nuclei obtained from three-dimensional NMR experiments were used to assign 95% of the backbone amino acid residues. Chemical shift data were analyzed using the TALOS+ program to detect the secondary structure elements and to estimate 1H-15N bond orientational order parameters. Our analysis leads to three important conclusions: the tertiary structure of OEP16 involves four transMembrane helices connected by flexible loop regions, it is likely to be monomeric in SDS micelles, and the ligand binding sites could be located. The α-helix content calculated in this method (55%) fits well with previous measurements (50%) [1]. Our 15N relaxation data supports the predicted secondary structure and helps in estimating the isotropic global rotational correlation time (τm) [2]. The value of τm indicates that OEP16 is a monomer in SDS micelles. The 2H exchange data revealed solvent exposed regions in the protein that aid in understanding the tertiary structural folding of the protein. The present study provides a basis for a high resolution structural determination of this Membrane protein in future NMR experiments. [1] D. Ni et al. (2011) Protein Expression Purif. 80, 157-168. [2] M. F. Brown (1982) J. Chem. Phys. 77, 1576-1599.