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1.
Proc Natl Acad Sci U S A ; 116(43): 21900-21906, 2019 10 22.
Artigo em Inglês | MEDLINE | ID: mdl-31591197

RESUMO

In plants, algae, and some photosynthetic bacteria, the ElectroChromic Shift (ECS) of photosynthetic pigments, which senses the electric field across photosynthetic membranes, is widely used to quantify the activity of the photosynthetic chain. In cyanobacteria, ECS signals have never been used for physiological studies, although they can provide a unique tool to study the architecture and function of the respiratory and photosynthetic electron transfer chains, entangled in the thylakoid membranes. Here, we identified bona fide ECS signals, likely corresponding to carotenoid band shifts, in the model cyanobacteria Synechococcus elongatus PCC7942 and Synechocystis sp. PCC6803. These band shifts, most likely originating from pigments located in photosystem I, have highly similar spectra in the 2 species and can be best measured as the difference between the absorption changes at 500 to 505 nm and the ones at 480 to 485 nm. These signals respond linearly to the electric field and display the basic kinetic features of ECS as characterized in other organisms. We demonstrate that these probes are an ideal tool to study photosynthetic physiology in vivo, e.g., the fraction of PSI centers that are prebound by plastocyanin/cytochrome c 6 in darkness (about 60% in both cyanobacteria, in our experiments), the conductivity of the thylakoid membrane (largely reflecting the activity of the ATP synthase), or the steady-state rates of the photosynthetic electron transport pathways.


Assuntos
Synechococcus/metabolismo , Tilacoides/metabolismo , Transporte de Elétrons , Eletrofisiologia , Potenciais da Membrana , Fotossíntese , Complexo de Proteína do Fotossistema I/metabolismo , Plastocianina/metabolismo
2.
Photosynth Res ; 142(3): 307-319, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31482263

RESUMO

A kinetic-LED-array-spectrophotometer (Klas) was recently developed for measuring in vivo redox changes of P700, plastocyanin (PCy), and ferredoxin (Fd) in the near-infrared (NIR). This spectrophotometer is used in the present work for in vitro light-induced measurements with various combinations of photosystem I (PSI) from tobacco and two different cyanobacteria, spinach plastocyanin, cyanobacterial cytochrome c6 (cyt. c6), and Fd. It is shown that cyt. c6 oxidation contributes to the NIR absorption changes. The reduction of (FAFB), the terminal electron acceptor of PSI, was also observed and the shape of the (FAFB) NIR difference spectrum is similar to that of Fd. The NIR difference spectra of the electron-transfer cofactors were compared between different organisms and to those previously measured in vivo, whereas the relative absorption coefficients of all cofactors were determined by using single PSI turnover conditions. Thus, the (840 nm minus 965 nm) extinction coefficients of the light-induced species (oxidized minus reduced for PC and cyt. c6, reduced minus oxidized for (FAFB), and Fd) were determined with values of 0.207 ± 0.004, - 0.033 ± 0.006, - 0.036 ± 0.008, and - 0.021 ± 0.005 for PCy, cyt. c6, (FAFB) (single reduction), and Fd, respectively, by taking a reference value of + 1 for P700+. The fact that the NIR P700 coefficient is larger than that of PCy and much larger than that of other contributing species, combined with the observed variability in the NIR P700 spectral shape, emphasizes that deconvolution of NIR signals into different components requires a very precise determination of the P700 spectrum.


Assuntos
Proteínas de Bactérias/química , Complexo de Proteína do Fotossistema I/metabolismo , Proteínas de Plantas/química , Espectroscopia de Luz Próxima ao Infravermelho/métodos , Proteínas de Bactérias/metabolismo , Citocromos c6/química , Citocromos c6/metabolismo , Transporte de Elétrons , Ferredoxinas/metabolismo , Oxirredução , Complexo de Proteína do Fotossistema I/química , Proteínas de Plantas/metabolismo , Plastocianina/química , Plastocianina/metabolismo , Espectrofotometria Ultravioleta , Espectroscopia de Luz Próxima ao Infravermelho/instrumentação , Spinacia oleracea/química , Synechocystis/química , Tabaco/química
3.
Nat Plants ; 5(6): 626-636, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31182847

RESUMO

In plants and green algae, the core of photosystem I (PSI) is surrounded by a peripheral antenna system consisting of light-harvesting complex I (LHCI). Here we report the cryo-electron microscopic structure of the PSI-LHCI supercomplex from the green alga Chlamydomonas reinhardtii. The structure reveals that eight Lhca proteins form two tetrameric LHCI belts attached to the PsaF side while the other two Lhca proteins form an additional Lhca2/Lhca9 heterodimer attached to the opposite side. The spatial arrangement of light-harvesting pigments reveals that Chlorophylls b are more abundant in the outer LHCI belt than in the inner LHCI belt and are absent from the core, thereby providing the downhill energy transfer pathways to the PSI core. PSI-LHCI is complexed with a plastocyanin on the patch of lysine residues of PsaF at the luminal side. The assembly provides a structural basis for understanding the mechanism of light-harvesting, excitation energy transfer of the PSI-LHCI supercomplex and electron transfer with plastocyanin.


Assuntos
Chlamydomonas reinhardtii/ultraestrutura , Complexos de Proteínas Captadores de Luz/ultraestrutura , Complexo de Proteína do Fotossistema I/química , Complexo de Proteína do Fotossistema I/ultraestrutura , Transferência de Energia , Proteínas de Membrana/química , Modelos Moleculares , Plastocianina/química , Conformação Proteica , Especificidade da Espécie
4.
Biochim Biophys Acta Bioenerg ; 1860(7): 591-599, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31247170

RESUMO

Small diffusible redox proteins play a ubiquitous role in bioenergetic systems, facilitating electron transfer (ET) between membrane bound complexes. Sustaining high ET turnover rates requires that the association between extrinsic and membrane-bound partners is highly specific, yet also sufficiently weak to promote rapid post-ET separation. In oxygenic photosynthesis the small soluble electron carrier protein plastocyanin (Pc) shuttles electrons between the membrane integral cytochrome b6f (cytb6f) and photosystem I (PSI) complexes. Here we use peak-force quantitative nanomechanical mapping (PF-QNM) atomic force microscopy (AFM) to quantify the dynamic forces involved in transient interactions between cognate ET partners. An AFM probe functionalised with Pc molecules is brought into contact with cytb6f complexes, immobilised on a planar silicon surface. PF-QNM interrogates the unbinding force of the cytb6f-Pc interactions at the single molecule level with picoNewton force resolution and on a time scale comparable to the ET time in vivo (ca. 120 µs). Using this approach, we show that although the unbinding force remains unchanged the interaction frequency increases over five-fold when Pc and cytb6f are in opposite redox states, so complementary charges on the cytb6f and Pc cofactors likely contribute to the electrostatic forces that initiate formation of the ET complex. These results suggest that formation of the docking interface is under redox state control, which lowers the probability of unproductive encounters between Pc and cytb6f molecules in the same redox state, ensuring the efficiency and directionality of this central reaction in the 'Z-scheme' of photosynthetic ET.


Assuntos
Complexo Citocromos b6f/metabolismo , Plastocianina/metabolismo , Análise de Célula Única/métodos , Spinacia oleracea/metabolismo , Complexo Citocromos b6f/química , Transporte de Elétrons , Oxirredução , Fotossíntese , Plastocianina/química , Ligação Proteica , Conformação Proteica
5.
J Phys Chem Lett ; 10(11): 2938-2943, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31074620

RESUMO

Alignment of molecules through electric fields minimizes the averaging over orientations, e.g., in single-particle-imaging experiments. The response of molecules to external ac electric fields is governed by their polarizability tensor, which is usually calculated using quantum chemistry methods. These methods are not feasible for large molecules. Here, we calculate the polarizability tensor of proteins using a regression model that correlates the polarizabilities of the 20 amino acids with perfect conductors of the same shape. The dielectric constant of the molecules could be estimated from the slope of the regression line based on the Clausius-Mossotti equation. We benchmark our predictions against the quantum chemistry results for the Trp cagemini protein and the measured dielectric constants of larger proteins. Our method has applications in computing laser alignment of macromolecules, for instance, benefiting single-particle imaging, as well as for estimation of the optical and electrostatic characteristics of proteins and other macromolecules.


Assuntos
Aminoácidos/química , Simulação por Computador , Anabaena variabilis/química , Cianobactérias/química , Inibidor da Ligação a Diazepam/química , Glutarredoxinas/química , Humanos , Plastocianina/química , Teoria Quântica , Análise de Regressão , Eletricidade Estática
6.
Physiol Plant ; 166(1): 320-335, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30740703

RESUMO

Mechanisms of the complex formation between plastocyanin and cytochrome f in higher plants (Spinacia oleracea and Brassica rapa), green microalgae Chlamydomonas reinhardtii and two species of cyanobacteria (Phormidium laminosum and Nostoc sp.) were investigated using combined Brownian and molecular dynamics simulations and hierarchical cluster analysis. In higher plants and green algae, electrostatic interactions force plastocyanin molecule close to the heme of cytochrome f. In the subsequent rotation of plastocyanin molecule around the point of electrostatic contact in the vicinity of cytochrome f, copper (Cu) atom approaches cytochrome heme forming a stable configuration where cytochrome f molecule behaves as a rather rigid body without conformational changes. In Nostoc plastocyanin molecule approaches cytochrome f in a different orientation (head-on) where the stabilization of the plastocyanin-cytochrome f complex is accompanied by the conformational changes of the G188E189D190 loop that stabilizes the whole complex. In cyanobacterium P. laminosum, electrostatic preorientation of the approaching molecules was not detected, thus indicating that random motions rather than long-range electrostatic interactions are responsible for the proper mutual orientation. We demonstrated that despite the structural similarity of the investigated electron transport proteins in different photosynthetic organisms, the complexity of molecular mechanisms of the complex formation increases in the following sequence: non-heterocystous cyanobacteria - heterocystous cyanobacteria - green algae - flowering plants.


Assuntos
Clorófitas/metabolismo , Cianobactérias/metabolismo , Citocromos f/metabolismo , Plastocianina/metabolismo , Transporte de Elétrons , Oxirredução , Complexo de Proteína do Fotossistema I/metabolismo , Espectrometria de Fluorescência
7.
Biochim Biophys Acta Bioenerg ; 1860(1): 60-68, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30414412

RESUMO

Cytochrome c6 is a soluble electron carrier, present in all known cyanobacteria, that has been replaced by plastocyanin in plants. Despite their high structural differences, both proteins have been reported to be isofunctional in cyanobacteria and green algae, acting as alternative electron carriers from the cytochrome b6-f complex to photosystem I or terminal oxidases. We have investigated the subcellular localization of both cytochrome c6 and plastocyanin in the heterocyst-forming cyanobacterium Anabaena sp. PCC 7120 grown in the presence of combined nitrogen and under diazotrophic conditions. Our studies conclude that cytochrome c6 is expressed at significant levels in heterocysts, even in the presence of copper, condition in which it is strongly repressed in vegetative cells. However, the copper-dependent regulation of plastocyanin is not altered in heterocysts. In addition, in heterocysts, cytochrome c6 has shown to be the main soluble electron carrier to cytochrome c oxidase-2 in respiration. A cytochrome c6 deletion mutant is unable to grow under diazotrophic conditions in the presence of copper, suggesting that cytochrome c6 plays an essential role in the physiology of heterocysts that cannot be covered by plastocyanin.


Assuntos
Anabaena/fisiologia , Respiração Celular , Citocromos c6/fisiologia , Fotossíntese , Cobre/farmacologia , Cianobactérias , Transporte de Elétrons , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Fixação de Nitrogênio , Plastocianina/fisiologia
8.
J Chem Phys ; 149(4): 045105, 2018 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-30068177

RESUMO

We investigate the effects of solvent specificities on the stability of the native structure (NS) of a protein on the basis of our free-energy function (FEF). We use CPB-bromodomain (CBP-BD) and apoplastocyanin (apoPC) as representatives of the protein universe and water, methanol, ethanol, and cyclohexane as solvents. The NSs of CBP-BD and apoPC consist of 66% α-helices and of 35% ß-sheets and 4% α-helices, respectively. In order to assess the structural stability of a given protein immersed in each solvent, we contrast the FEF of its NS against that of a number of artificially created, misfolded decoys possessing the same amino-acid sequence but significantly different topology and α-helix and ß-sheet contents. In the FEF, we compute the solvation entropy using the morphometric approach combined with the integral equation theories, and the change in electrostatic (ES) energy upon the folding is obtained by an explicit atomistic but simplified calculation. The ES energy change is represented by the break of protein-solvent hydrogen bonds (HBs), formation of protein intramolecular HBs, and recovery of solvent-solvent HBs. Protein-solvent and solvent-solvent HBs are absent in cyclohexane. We are thus able to separately evaluate the contributions to the structural stability from the entropic and energetic components. We find that for both CBP-BD and apoPC, the energetic component dominates in methanol, ethanol, and cyclohexane, with the most stable structures in these solvents sharing the same characteristics described as an association of α-helices. In particular, those in the two alcohols are identical. In water, the entropic component is as strong as or even stronger than the energetic one, with a large gain of translational, configurational entropy of water becoming crucially important so that the relative contents of α-helix and ß-sheet and the content of total secondary structures are carefully selected to achieve sufficiently close packing of side chains. If the energetic component is excluded for a protein in water, the priority is given to closest side-chain packing, giving rise to the formation of a structure with very low α-helix and ß-sheet contents. Our analysis, which requires minimal computational effort, can be applied to any protein immersed in any solvent and provides robust predictions that are quite consistent with the experimental observations for proteins in different solvent environments, thus paving the way toward a more detailed understanding of the folding process.


Assuntos
Apoproteínas/química , Plastocianina/química , Solventes/química , Ligação de Hidrogênio , Conformação Proteica , Domínios Proteicos , Eletricidade Estática
9.
Bioelectrochemistry ; 124: 127-132, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30029033

RESUMO

Increasing the thermal stability of immobilized proteins is a motivating goal for improving the performance of electrochemical biodevices. In this work, we propose the immobilization of crosslinked plastocyanin from the thermophilic cyanobacterium Phormidium laminosum by simultaneous incubation of a mixture of plastocyanin and the coupling reagents. The thermal stability of the so built covalently immobilized protein films has been assessed by cyclic voltammetry in the 0-90 °C temperature range and has been compared to that of physisorbed films. It is shown that the protein loss along a thermal cycle is significantly reduced in the case of the crosslinked films, whose redox properties remain unaltered along a cyclic heating-cooling thermal scan, and can withstand the contact with 70 °C solutions for four hours. Comparison of thermal unfolding curves obtained by circular dichroism spectroscopy of both free and crosslinked protein confirms the improved thermic resistance of the crosslinked plastocyanin. Notably, the electron transfer thermodynamics of physisorbed and crosslinked plastocyanin films are quite similar, suggesting that the formation of intra- and inter-protein amide bonds do not affect the integrity and functionality of the copper redox centers. UV-Vis absorption and circular dichroism measurements corroborate that protein crosslinking does not alter the coordination geometry of the metal center.


Assuntos
Cianobactérias/química , Eletrodos , Ouro/química , Plastocianina/química , Proteínas/química , Dicroísmo Circular , Cobre/química , Reagentes para Ligações Cruzadas/química , Temperatura Alta , Oxirredução , Estabilidade Proteica , Espectrofotometria Ultravioleta , Termodinâmica
10.
Photosynth Res ; 137(2): 281-293, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29594952

RESUMO

The absolute amount of plastocyanin (PC), ferredoxin-NADP+-oxidoreductase (FNR), hydrogenase (HYDA1), and ferredoxin 5 (FDX5) were quantified in aerobic and anaerobic Chlamydomonas reinhardtii whole cells using purified (recombinant) proteins as internal standards in a mass spectrometric approach. Quantified protein amounts were related to the estimated amount of PSI. The ratios of PC to FNR to HYDA1 to FDX5 in aerobic cells were determined to be 1.4:1.2:0.003:0. In anaerobic cells, the ratios changed to 1.1:1.3:0.019:0.027 (PC:FNR:HYDA1:FDX5). Employing sodium dithionite and methyl viologen as electron donors, the specific activity of hydrogenase in whole cells was calculated to be 382 ± 96.5 µmolH2 min-1 mg-1. Importantly, these data reveal an about 70-fold lower abundance of HYDA1 compared to FNR. Despite this great disproportion between both proteins, which might further enhance the competition for electrons, the alga is capable of hydrogen production under anaerobic conditions, thus pointing to an efficient channeling mechanism of electrons from FDX1 to the HYDA1.


Assuntos
Chlamydomonas reinhardtii/metabolismo , Ferredoxinas/metabolismo , Hidrogenase/metabolismo , Nitrito Redutases/metabolismo , Fotossíntese/fisiologia , Plastocianina/metabolismo , Aerobiose , Sequência de Aminoácidos , Anaerobiose , Ferredoxinas/genética , Regulação da Expressão Gênica/fisiologia , Regulação Enzimológica da Expressão Gênica , Hidrogenase/genética , Nitrito Redutases/genética , Oxigênio/metabolismo , Oxigênio/farmacologia
11.
J Integr Plant Biol ; 60(4): 323-340, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29330900

RESUMO

The ability of a plant to produce grain, fruit, or forage depends ultimately on photosynthesis. There have been few attempts, however, to study microRNAs, which are a class of endogenous small RNAs post-transcriptionally programming gene expression, in relation to photosynthetic traits. We focused on miR408, one of the most conserved plant miRNAs, and overexpressed it in parallel in Arabidopsis, tobacco, and rice. The transgenic plants all exhibited increased copper content in the chloroplast, elevated abundance of plastocyanin, and an induction of photosynthetic genes. By means of gas exchange and optical spectroscopy analyses, we showed that higher expression of miR408 leads to enhanced photosynthesis through improving efficiency of irradiation utilization and the capacity for carbon dioxide fixation. Consequently, miR408 hyper-accumulating plants exhibited higher rate of vegetative growth. An enlargement of seed size was also observed in all three species overproducing miR408. Moreover, we conducted a 2-year-two-location field trial and observed miR408 overexpression in rice significantly increased yield, which was primarily attributed to an elevation in grain weight. Taken together, these results demonstrate that miR408 is a positive regulator of photosynthesis and that its genetic engineering is a promising route for enhancing photosynthetic performance and yield in diverse plants.


Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , MicroRNAs/metabolismo , Fotossíntese/genética , Sementes/crescimento & desenvolvimento , Sementes/genética , Cloroplastos/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Oryza/genética , Plantas Geneticamente Modificadas , Plastocianina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Tabaco/genética
12.
Plant Sci ; 268: 1-10, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29362078

RESUMO

Accumulating evidence indicates that plant plastocyanin is involved in copper homeostasis, yet the physiological relevance remains elusive. In this study, we found that a plastocyanin gene (SsPETE2) from euhalophyte Suaeda salsa possessed a novel antioxidant function, which was associated with the copper-chelating activity of SsPETE2. In S. salsa, expression of SsPETE2 increased in response to oxidative stress and ectopic expression of SsPETE2 in Arabidopsis enhanced the antioxidant ability of the transgenic plants. SsPETE2 bound Cu ion and alleviated formation of hydroxyl radicals in vitro. Accordingly, SsPETE2 expression lowered the free Cu content that was associated with reduced H2O2 level under oxidative stress. Arabidopsis pete1 and pete2 mutants showed ROS-sensitive phenotypes that could be restored by expression of SsPETE2 or AtPETEs. In addition, SsPETE2-expressing plants exhibited more potent tolerance to oxidative stress than plants overexpressing AtPETEs, likely owing to the stronger copper-binding activity of SsPETE2 than AtPETEs. Taken together, these results demonstrated that plant PETEs play a novel role in oxidative stress tolerance by regulating Cu homeostasis under stress conditions, and SsPETE2, as an efficient copper-chelating PETE, potentially could be used in crop genetic engineering.


Assuntos
Adaptação Fisiológica , Chenopodiaceae/genética , Chenopodiaceae/fisiologia , Expressão Ectópica do Gene , Estresse Oxidativo/genética , Proteínas de Plantas/metabolismo , Plastocianina/genética , Adaptação Fisiológica/efeitos dos fármacos , Antioxidantes/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Quelantes/farmacologia , Chenopodiaceae/efeitos dos fármacos , Cloroplastos/efeitos dos fármacos , Cloroplastos/metabolismo , Cobre/farmacologia , Desoxirribose/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacologia , Radical Hidroxila/metabolismo , Íons , Ferro/metabolismo , Simulação de Dinâmica Molecular , Mutação/genética , Estresse Oxidativo/efeitos dos fármacos , Paraquat/farmacologia , Fenótipo , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Plastocianina/metabolismo , Transporte Proteico/efeitos dos fármacos
13.
Proc Natl Acad Sci U S A ; 115(6): 1280-1285, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29358375

RESUMO

Oxidoreductases catalyze electron transfer reactions that ultimately provide the energy for life. A limited set of ancestral protein-metal modules are presumably the building blocks that evolved into this diverse protein family. However, the identity of these modules and their path to modern oxidoreductases is unknown. Using a comparative structural analysis approach, we identify a set of fundamental electron transfer modules that have evolved to form the extant oxidoreductases. Using transition metal-containing cofactors as fiducial markers, it is possible to cluster cofactor microenvironments into as few as four major modules: bacterial ferredoxin, cytochrome c, symerythrin, and plastocyanin-type folds. From structural alignments, it is challenging to ascertain whether modules evolved from a single common ancestor (homology) or arose by independent convergence on a limited set of structural forms (analogy). Additional insight into common origins is contained in the spatial adjacency network (SPAN), which is based on proximity of modules in oxidoreductases containing multiple cofactor electron transfer chains. Electron transfer chains within complex modern oxidoreductases likely evolved through repeated duplication and diversification of ancient modular units that arose in the Archean eon.


Assuntos
Coenzimas/metabolismo , Evolução Molecular , Oxirredutases/química , Oxirredutases/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Coenzimas/química , Citocromos c/química , Citocromos c/metabolismo , Transporte de Elétrons , Ferredoxinas/química , Ferredoxinas/metabolismo , Metais/química , Metais/metabolismo , Modelos Moleculares , Plastocianina/química , Plastocianina/metabolismo , Conformação Proteica , Homologia Estrutural de Proteína
14.
Plant Physiol ; 175(3): 1175-1185, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28904074

RESUMO

Increasing grain yield is the most important object of crop breeding. Here, we report that the elevated expression of a conserved microRNA, OsmiR408, could positively regulate grain yield in rice (Oryza sativa) by increasing panicle branches and grain number. We further showed that OsmiR408 regulates grain yield by down-regulating its downstream target, OsUCL8, which is an uclacyanin (UCL) gene of the phytocyanin family. The knock down or knock out of OsUCL8 also increases grain yield, while the overexpression of OsUCL8 results in an opposite phenotype. Spatial and temporal expression analyses showed that OsUCL8 was highly expressed in pistils, young panicles, developing seeds, and inflorescence meristem and was nearly complementary to that of OsmiR408. Interestingly, the OsUCL8 protein was localized to the cytoplasm, distinct from a majority of phytocyanins, which localize to the plasma membrane. Further studies revealed that the cleavage of OsUCL8 by miR408 affects copper homeostasis in the plant cell, which, in turn, affects the abundance of plastocyanin proteins and photosynthesis in rice. To our knowledge, this is the first report of the effects of miR408-OsUCL8 in regulating rice photosynthesis and grain yield. Our study further broadens the perspective of microRNAs and UCLs and provides important information for breeding high-yielding crops through genetic engineering.


Assuntos
MicroRNAs/metabolismo , Oryza/genética , Oryza/fisiologia , Fotossíntese , Plastocianina/metabolismo , Sementes/genética , Sementes/fisiologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , MicroRNAs/genética , Oryza/anatomia & histologia , Fenótipo , Fotossíntese/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Interferência de RNA
15.
Photosynth Res ; 134(3): 343-360, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28497192

RESUMO

Properties and performance of the recently introduced Dual/KLAS-NIR spectrophotometer for simultaneous measurements of ferredoxin (Fd), P700, and plastocyanin (PC) redox changes, together with whole leaf chlorophyll a (Chl) fluorescence (emission >760, 540 nm excitation) are outlined. Spectral information on in vivo Fd, P700, and PC in the near-infrared region (NIR, 780-1000 nm) is presented, on which the new approach is based. Examples of application focus on dark-light and light-dark transitions, where maximal redox changes of Fd occur. After dark-adaptation, Fd reduction induced by moderate light parallels the Kautsky effect of Chl fluorescence induction. Both signals are affected analogously by removal of O2. A rapid type of Fd reoxidation, observed after a short pulse of light before light activation of linear electron transport (LET), is more pronounced in C4 compared to C3 leaves and interpreted to reflect cyclic PS I (CET). Light activation of LET, as assessed via the rate of Fd reoxidation after short light pulses, occurs at very low intensities and is slowly reversed (half-time ca. 20 min). Illumination with strong far-red light (FR, 740 nm) reveals two fractions of PS I, PS I (LET), and PS I (CET), differing in the rates of Fd reoxidation upon FR-off and the apparent equilibrium constants between P700 and PC. Parallel information on oxidation of Fd and reduction of P700 plus PC proves essential for identification of CET. Comparison of maize (C4) with sunflower and ivy (C3) responses leads to the conclusion that segregation of two types of PS I may not only exist in C4 (mesophyll and bundle sheath cells), but also in C3 photosynthesis (grana margins plus end membranes and stroma lamellae).


Assuntos
Ferredoxinas/metabolismo , Luz , Folhas de Planta/metabolismo , Plantas/metabolismo , Plastocianina/metabolismo , Elétrons , Fluorescência , Cinética , Oxirredução/efeitos da radiação , Complexo de Proteína do Fotossistema I/metabolismo , Folhas de Planta/efeitos da radiação , Fatores de Tempo
16.
Physiol Plant ; 161(1): 88-96, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28369912

RESUMO

A model of electron transport from cytochrome f to photosystem I mediated by plastocyanin was designed on the basis of the multiparticle Brownian dynamics method. The model combines events which occur over a wide time range, including protein diffusion along the thylakoid membrane, long-distance interactions between proteins, formation of a multiprotein complex, electron transfer within a complex and complex dissociation. Results of the modeling were compared with the experimental kinetics measured in chloroplast thylakoids. Computer simulation demonstrated that the complex interior of the photosynthetic membrane, electrostatic interactions and Brownian diffusion provide physical conditions for the directed electron flow along the photosynthetic electron transport chain.


Assuntos
Simulação por Computador , Complexo Citocromos b6f/metabolismo , Modelos Moleculares , Complexo de Proteína do Fotossistema I/metabolismo , Plastocianina/metabolismo , Clorofila/metabolismo , Transporte de Elétrons , Cinética , Modelos Biológicos , Oxirredução , Eletricidade Estática , Fatores de Tempo
17.
J Exp Bot ; 68(5): 1137-1155, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28180288

RESUMO

PsaI is the only subunit of PSI whose precise physiological function has not yet been elucidated in higher plants. While PsaI is involved in PSI trimerization in cyanobacteria, trimerization was lost during the evolution of the eukaryotic PSI, and the entire PsaI side of PSI underwent major structural remodelling to allow for binding of light harvesting complex II antenna proteins during state transitions. Here, we have generated a tobacco (Nicotiana tabacum) knockout mutant of the plastid-encoded psaI gene. We show that PsaI is not required for the redox reactions of PSI. Neither plastocyanin oxidation nor the processes at the PSI acceptor side are impaired in the mutant, and both linear and cyclic electron flux rates are unaltered. The PSI antenna cross section is unaffected, state transitions function normally, and binding of other PSI subunits to the reaction centre is not compromised. Under a wide range of growth conditions, the mutants are phenotypically and physiologically indistinguishable from wild-type tobacco. However, in response to high-light and chilling stress, and especially during leaf senescence, PSI content is reduced in the mutants, indicating that the I-subunit plays a role in stabilizing PSI complexes.


Assuntos
Folhas de Planta/fisiologia , Proteínas de Plantas/genética , Tabaco/genética , Oxirredução , Complexo de Proteína do Fotossistema I/genética , Complexo de Proteína do Fotossistema I/metabolismo , Proteínas de Plantas/metabolismo , Plastídeos/metabolismo , Plastocianina/metabolismo , Tabaco/metabolismo
18.
Plant Cell Physiol ; 58(2): 256-265, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28007969

RESUMO

All known cyanobacteria contain Cyt c6, a small soluble electron carrier protein whose main function is to transfer electrons from the Cyt b6f complex to PSI, although it is also involved in respiration. We have previously described a second isoform of this protein, the Cyt c6-like, whose function remains unknown. Here we describe a third isoform of Cyt c6 (here called Cytc6-3), which is only found in heterocyst-forming filamentous cyanobacteria. Cyt c6-3 is expressed in vegetative cells but is specifically repressed in heterocysts cells under diazotrophic growth conditions. Although there is a close structural similarity between Cyt c6-3 and Cyt c6 related to the general protein folding, Cyt c6-3 presents differential electrostatic surface features as compared with Cyt c6, its expression is not copper dependent and has a low reactivity towards PSI. According to the different expression pattern, functional reactivity and structural properties, Cyt c6-3 has to play an as yet to be defined regulatory role related to heterocyst differentiation.


Assuntos
Proteínas de Bactérias/metabolismo , Cianobactérias/metabolismo , Isoformas de Proteínas/metabolismo , Transporte de Elétrons/fisiologia , Fotossíntese/fisiologia , Plastocianina/metabolismo
19.
J Comput Chem ; 38(16): 1431-1437, 2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27859435

RESUMO

Plastocyanin is a copper containing protein that is involved in the electron transfer process in photosynthetic organisms. The active site of plastocyanin is described as an entatic state whereby its structure represents a compromise between the structures favored by the oxidized and reduced forms. In this study, the nature of the entatic state is investigated through density functional theory-based hybrid quantum mechanics/molecular mechanics (QM/MM) molecular dynamics simulations. The strain energy is computed to be 12.8 kcal/mol and 14.5 kcal/mol for the oxidized and reduced forms of the protein, indicating that the active site has an intermediate structure. It is shown that the energy gap between the oxidized and reduced forms varies significantly with the fluctuations in the structure of the active site at room temperature. An accurate determination of the reorganization energy requires averaging over conformation and a large region of the protein around the active site to be treated at the quantum mechanical level. © 2016 The Authors. Journal of Computational Chemistry Published by Wiley Periodicals, Inc.


Assuntos
Simulação de Dinâmica Molecular , Plastocianina/química , Domínio Catalítico , Transporte de Elétrons , Oxirredução , Conformação Proteica , Teoria Quântica , Termodinâmica
20.
FEBS Lett ; 590(20): 3639-3648, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27685247

RESUMO

Plastocyanin (petE) plays an essential role in photosynthesis as an electron carrier between cytochrome b6 f and photosystem I, and in some cyanobacteria it can be replaced by the haem-containing protein, cytochrome c6 (petJ). In Synechocystis sp. PCC 6803, transcription of petE and petJ is activated and repressed, respectively, by Cu. Here, we show that Ni can act similarly to Cu in inducing petE and repressing petJ, thus leading to a partial switch between cytochrome c6 and plastocyanin. Transcription of these genes is only altered by Ni in Cu-depleted medium, and none of the Ni-dependent transcription factors described in Synechocystis, NrsR and InrS seem to be involved in this regulation. Finally, we show that plastocyanin is essential for growth under conditions of excess Ni.


Assuntos
Citocromos c6/genética , Níquel/metabolismo , Plastocianina/genética , Synechocystis/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Citocromos c6/metabolismo , Regulação Bacteriana da Expressão Gênica , Plastocianina/metabolismo , Synechocystis/genética , Synechocystis/metabolismo , Transcrição Genética
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