ferredoxin-NADP Reductase Research Articles

Intensifying Electron Utilization by Surface-Anchored Rh Complex for Enhanced Nicotinamide Cofactor Regeneration and Photoenzymatic CO2 Reduction.

Solar-driven photocatalytic regeneration of cofactors, including reduced nicotinamide adenine dinucleotide (NADH), reduced nicotinamide adenine dinucleotide phosphate (NADPH), and reduced flavin adenine dinucleotide (FADH2), could ensure the sustainable energy supply of enzymatic reactions catalyzed by oxidoreductases for the efficient synthesis of chemicals. However, the elevation of cofactor regeneration efficiency is severely hindered by the inefficient utilization of electrons transferred on the surface of photocatalysts. Inspired by the phenomenon of ferredoxin-NADP+ reductase (FNR) anchoring on thylakoid membrane, herein, a homogeneous catalyst of rhodium (Rh) complex, [Cp∗Rh(bpy)H2O]2+, was anchored on polymeric carbon nitride (PCN) mediated by a tannic acid/polyethyleneimine (TA/PEI) adhesive layer, acquiring PCN@TA/PEI-Rh core@shell photocatalyst. Illuminated by visible light, electrons were excited from the PCN core, then transferred through the TA/PEI shell, and finally captured by the surface-anchored Rh for instant utilization during the regeneration of NADH. The TA/PEI-Rh shell could facilitate the electron transfer from the PCN core and, more importantly, achieved ~1.3-fold elevation of electron utilization efficiency compared with PCN. Accordingly, the PCN@TA/PEI-Rh afforded the NADH regeneration efficiency of 37.8% after 20 min reaction under LED light (405 nm) illumination, over 1.5 times higher than PCN with free Rh. Coupling of the NADH regeneration system with formate dehydrogenase achieved continuous production of formate from carbon dioxide (CO2). Our study may provide a generic and effective strategy to elevate the catalytic efficiency of a photocatalyst through intensifying the electron utilization.

Progress in Scaling up and Streamlining a Nanoconfined, Enzyme-Catalyzed Electrochemical Nicotinamide Recycling System for Biocatalytic Synthesis.

An electrochemically driven nicotinamide recycling system, referred to as the ‘electrochemical leaf’ has unique attributes that may suit it to the small‐scale industrial synthesis of high‐value chemicals. A complete enzyme cascade can be immobilized within the channels of a nanoporous electrode, allowing complex reactions to be energized, controlled and monitored continuously in real time. The electrode is easily prepared by depositing commercially available indium tin oxide (ITO) nanoparticles on a Ti support, resulting in a network of nanopores into which enzymes enter and bind. One of the enzymes is the photosynthetic flavoenzyme, ferredoxin NADP+ reductase (FNR), which catalyzes the quasi‐reversible electrochemical recycling of NADP(H) and serves as the transducer. The second enzyme is any NADP(H)‐dependent dehydrogenase of choice, and further enzymes can be added to build elaborate cascades that are driven in either oxidation or reduction directions through the rapid recycling of NADP(H) within the pores. In this Article, we describe the measurement of key enzyme/cofactor parameters and an essentially linear scale‐up from an analytical scale 4 mL reactor with a 14 cm2 electrode to a 500 mL reactor with a 500 cm2 electrode. We discuss the advantages (energization, continuous monitoring that can be linked to a computer, natural enzyme immobilization, low costs of electrodes and low cofactor requirements) and challenges to be addressed (optimizing minimal use of enzyme applied to the electrode).

Nitric oxide and hydrogen sulfide modulate the NADPH-generating enzymatic system in higher plants.

Nitric oxide (NO) and hydrogen sulfide (H2S) are two key molecules in plant cells that participate, directly or indirectly, as regulators of protein functions through derived post-translational modifications, mainly tyrosine nitration, S-nitrosation, and persulfidation. These post-translational modifications allow the participation of both NO and H2S signal molecules in a wide range of cellular processes either physiological or under stressful circumstances. NADPH participates in cellular redox status and it is a key cofactor necessary for cell growth and development. It is involved in significant biochemical routes such as fatty acid, carotenoid and proline biosynthesis, and the shikimate pathway, as well as in cellular detoxification processes including the ascorbate-glutathione cycle, the NADPH-dependent thioredoxin reductase (NTR), or the superoxide-generating NADPH oxidase. Plant cells have diverse mechanisms to generate NADPH by a group of NADP-dependent oxidoreductases including ferredoxin-NADP reductase (FNR), NADP-glyceraldehyde-3-phosphate dehydrogenase (NADP-GAPDH), NADP-dependent malic enzyme (NADP-ME), NADP-dependent isocitrate dehydrogenase (NADP-ICDH), and both enzymes of the oxidative pentose phosphate pathway, designated as glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH). These enzymes consist of different isozymes located in diverse subcellular compartments (chloroplasts, cytosol, mitochondria, and peroxisomes) which contribute to the NAPDH cellular pool. We provide a comprehensive overview of how post-translational modifications promoted by NO (tyrosine nitration and S-nitrosation), H2S (persulfidation), and glutathione (glutathionylation), affect the cellular redox status through regulation of the NADP-dependent dehydrogenases.

A novel chloroplast super-complex consisting of the ATP synthase and photosystem I reaction center.

Several ‘super-complexes’ of individual hetero-oligomeric membrane protein complexes, whose function is to facilitate intra-membrane electron and proton transfer and harvesting of light energy, have been previously characterized in the mitochondrial cristae and chloroplast thylakoid membranes. We report the presence of an intra-membrane super-complex dominated by the ATP-synthase, photosystem I (PSI) reaction-center complex and the ferredoxin-NADP+ Reductase (FNR) in the thylakoid membrane. The presence of the super-complex has been documented by mass spectrometry, clear-native PAGE and Western Blot analyses. This is the first documented presence of ATP synthase in a super-complex with the PSI reaction-center located in the non-appressed stromal domain of the thylakoid membrane.

Transcriptomic and network analyses reveal distinct nitrate responses in light and dark in rice leaves (Oryza sativa Indica var. Panvel1)

Nitrate (N) response is modulated by light, but not understood from a genome-wide perspective. Comparative transcriptomic analyses of nitrate response in light-grown and etiolated rice leaves revealed 303 and 249 differentially expressed genes (DEGs) respectively. A majority of them were exclusive to light (270) or dark (216) condition, whereas 33 DEGs were common. The latter may constitute response to N signaling regardless of light. Functional annotation and pathway enrichment analyses of the DEGs showed that nitrate primarily modulates conserved N signaling and metabolism in light, whereas oxidation–reduction processes, pentose-phosphate shunt, starch-, sucrose- and glycerolipid-metabolisms in the dark. Differential N-regulation of these pathways by light could be attributed to the involvement of distinctive sets of transporters, transcription factors, enriched cis-acting motifs in the promoters of DEGs as well as differential modulation of N-responsive transcriptional regulatory networks in light and dark. Sub-clustering of DEGs-associated protein–protein interaction network constructed using experimentally validated interactors revealed that nitrate regulates a molecular complex consisting of nitrite reductase, ferredoxin-NADP reductase and ferredoxin. This complex is associated with flowering time, revealing a meeting point for N-regulation of N-response and N-use efficiency. Together, our results provide novel insights into distinct pathways of N-signaling in light and dark conditions.

Mimicking Functions of Native Enzymes or Photosynthetic Reaction Centers by Nucleoapzymes and Photonucleoapzymes.

The covalent linkage of catalytic units to aptamer sequence-specific nucleic acids exhibiting selective binding affinities for substrates leads to functional scaffolds mimicking native enzymes, nucleoapzymes. The binding of the substrates to the aptamer and their structural orientation with respect to the catalytic units duplicate the functions of the active center of enzymes. The possibility of linking the catalytic sites directly, or through spacer units, to the 5′-end, 3′-end, and middle positions of the aptamers allows the design of nucleoapzyme libraries, revealing structure–functions diversities, and these can be modeled by molecular dynamics simulations. Catalytic sites integrated into nucleoapzymes include DNAzymes, transition metal complexes, and organic ligands. Catalytic transformations driven by nucleoapzymes are exemplified by the oxidation of dopamine or l-arginine, hydroxylation of tyrosine to l-DOPA, hydrolysis of ATP, and cholic acid-modified esters. The covalent linkage of photosensitizers to the tyrosinamide aptamer leads to a photonucleoapzyme scaffold that binds the N-methyl-N′-(3-aminopropane)-4,4′-bipyridinium-functionalized tyrosinamide to the aptamer. By linking the photosensitizer directly, or through a spacer bridge to the 5′-end or 3′-end of the aptamer, we demonstrate a library of supramolecular photosensitizer/electron acceptor photonucleoapzymes mimicking the functions of photosystem I in the photosynthetic apparatus. The photonucleoapzymes catalyze the photoinduced generation of NADPH, in the presence of ferredoxin-NADP+-reductase (FNR), or the photoinduced H2 evolution catalyzed by Pt nanoparticles. The future prospects of nucleoapzymes and photonucleoapzymes are discussed.

Proteomic analyses unraveling water stress response in two Eucalyptus species originating from contrasting environments for aridity.

Eucalyptus are widely cultivated in several regions of the world due to their adaptability to different climatic conditions and amenable to tree breeding programs. With changes in environmental conditions pointing to an increase in aridity in many areas of the globe, the demand for genetic materials that adapt to this situation is required. Therefore, the aim of this work was to identify contrasting differences between two Eucalyptus species under water stress through the identification of differentially abundant proteins. For this, total protein extraction was proceeded from leaves of both species maintained at 40 and 80% of field capacity (FC). The 80% FC water regime was considered as the control and the 40% FC, severe water stress. The proteins were separated by 2-DE with subsequent identification of those differentially abundant by liquid nanocromatography coupled to high resolution MS (Q-Exactive). Comparative proteomics allowed to identify four proteins (ATP synthase gamma and alpha, glutamine synthetase and a vacuolar protein) that were more abundant in drought-tolerant species and simultaneously less abundant or unchanged in the drought- sensitive species, an uncharacterized protein found exclusively in plants under drought stress and also 10 proteins (plastid-lipid, ruBisCO activase, ruBisCO, protease ClpA, transketolase, isoflavone reductase, ferredoxin-NADP reductase, malate dehydrogenase, aminobutyrate transaminase and sedoheptulose-1-bisphosphatase) induced exclusively in the drought-tolerant species in response to water stress. These results suggest that such proteins may play a crucial role as potential markers of water stress tolerance through the identification of species-specific proteins, and future targets for genetic engineering.

Structural basis for plant lutein biosynthesis from α-carotene

Two cytochrome P450 enzymes, CYP97A3 and CYP97C1, catalyze hydroxylations of the β- and ε-rings of α-carotene to produce lutein. Chirality is introduced at the C-3 atom of both rings, and the reactions are both pro-3R-stereospecific. We determined the crystal structures of CYP97A3 in substrate-free and complex forms with a nonnatural substrate and the structure of CYP97C1 in a detergent-bound form. The structures of CYP97A3 in different states show the substrate channel and the structure of CYP97C1 bound with octylthioglucoside confirms the binding site for the carotenoid substrate. Biochemical assays confirm that the ferredoxin-NADP+ reductase (FNR)-ferredoxin pair is used as the redox partner. Details of the pro-3R stereospecificity are revealed in the retinal-bound CYP97A3 structure. Further analysis indicates that the CYP97B clan bears similarity to the β-ring-specific CYP97A clan. Overall, our research describes the molecular basis for the last steps of lutein biosynthesis.

Special issue in honour of Prof. Reto J. Strasser - Effect of AtLFNR1 deficiency on chlorophyll a fluorescence rise kinetics OJIP of Arabidopsis

Leaf-type ferredoxin-NADP(H) oxidoreductase, encoded by AtLFNR1 gene in Arabidopsis, extensively exists in chloroplast stroma and thylakoids and is responsible for the reduction of NADP+ to NADPH in PSI. To investigate the relationship between the characteristics of chlorophyll a fluorescence transients OJIP and the function of AtLFNR1 protein, the fluorescence rise kinetics OJIP curves of AtLFNR1 mutant leaves were determined. It is observed that AtLFNR1 mutant has a lower level of variable fluorescence intensity relative to wild type, especially the J-step and the IP-phase. The JIP-test analysis suggests that the loss of AtLFNR1 function decreases clearly the oxidation rate of plastocyanin as well as PSI reaction center, damages slightly PSII antenna size and pigment assemblies, but stimulates distinctly the photosynthetic activity of PSII. Such multiple effects contribute to the change of shape characteristics of the OJIP transient curve in AtLFNR1 mutant.

Molecular basis of ferredoxin:NADP(+) reductase interactions with FNR binding domains from TROL and Tic62 proteins

The final step of photosynthetic electron transfer from ferredoxin to NADP+ is catalysed by ferredoxin: NADP+ oxidoreductase (FNR). Localization of the FNR on the chloroplastic membrane is attained both, by the dynamic interaction via thylakoid rhodanese-like protein (TROL) and with the lasting interaction with the Tic62 protein. In such interactions, FNR forms a homodimeric complex in the presence of the FNR interacting domain from either TROL or Tic62 protein. Starting from crystal structure of FNR homodimeric complex, various systems consisting of the FNR proteins and/or the FNR binding domains from TROL and Tic62 proteins at the different pH were subjected to molecular dynamics simulations. Results of simulations enabled identification of the most important regions of non-covalent interactions between involved proteins and their detailed mapping. In silico mutations were introduced and simulated to investigate their influence to the structural properties, dynamics and stability of the protein complex. The interface regions between the FNR proteins, consisting mostly of hydrophobic interactions, were identified and shown to be under strong influence of the ITEPs conserved prolines and serines (PPSS region). Five functionally important hydrogen bond networks that span over the major structural elements of the involved proteins were identified and characterized. Three of them show pH dependence and might serve as the pH regulated switches for conformational changes of the protein complex. Presented computational results provide strong base for in vitro experiments on TROL, Tic62 and FNR proteins that should enable understanding of their behavior in the fluctuating environmental conditions.

Modelling Photosynthesis with ZnII -Protoporphyrin All-DNA G-Quadruplex/Aptamer Scaffolds.

All-DNA scaffolds act as templates for the organization of photosystem I model systems. A series of DNA templates composed of ZnII -protoporphyrin IX (ZnII PPIX)-functionalized G-quadruplex conjugated to the 3'- or 5'-end of the tyrosinamide (TA) aptamer and ZnII PPIX/G-quadruplex linked to the 3'- and 5'-ends of the TA aptamer through a four-thymidine bridge. Effective photoinduced electron transfer (ET) from ZnII PPIX/G-quadruplex to bipyridinium-functionalized tyrosinamide, TA-MV2+ , bound to the TA aptamer units is demonstrated. The effectiveness of the primary ET quenching of ZnII PPIX/G-quadruplex by TA-MV2+ controls the efficiency of the generation of TA-MV+. . The photosystem-controlled formation of TA-MV+. by the different photosystems dictates the secondary activation of the ET cascade corresponding to the ferredoxin-NADP+ reductase (FNR)-catalysed reduction of NADP+ to NADPH by TA-MV+. , and the sequestered alcohol dehydrogenase catalysed reduction of acetophenone to 1-phenylethanol by NADPH.

Modelling Photosynthesis with ZnII‐Protoporphyrin All‐DNA G‐Quadruplex/Aptamer Scaffolds

AbstractAll‐DNA scaffolds act as templates for the organization of photosystem I model systems. A series of DNA templates composed of ZnII‐protoporphyrin IX (ZnIIPPIX)‐functionalized G‐quadruplex conjugated to the 3′‐ or 5′‐end of the tyrosinamide (TA) aptamer and ZnIIPPIX/G‐quadruplex linked to the 3′‐ and 5′‐ends of the TA aptamer through a four‐thymidine bridge. Effective photoinduced electron transfer (ET) from ZnIIPPIX/G‐quadruplex to bipyridinium‐functionalized tyrosinamide, TA‐MV2+, bound to the TA aptamer units is demonstrated. The effectiveness of the primary ET quenching of ZnIIPPIX/G‐quadruplex by TA‐MV2+ controls the efficiency of the generation of TA‐MV+.. The photosystem‐controlled formation of TA‐MV+. by the different photosystems dictates the secondary activation of the ET cascade corresponding to the ferredoxin‐NADP+ reductase (FNR)‐catalysed reduction of NADP+ to NADPH by TA‐MV+., and the sequestered alcohol dehydrogenase catalysed reduction of acetophenone to 1‐phenylethanol by NADPH.

Photochemistry and proteomics of ginger (Zingiber officinale Roscoe) under drought and shading

Drought has become an increasingly serious ecological problem that limits crop production. However, little is known about the response of ginger (Zingiber officinale Roscoe) to drought and shading, especially with respect to photosynthetic electron transport. Here, differential proteomics was used to study the response of ginger to four experimental treatments: control, drought, 50% shading, and the combination of 50% shading and drought. Proteomic analysis suggested that ginger increased cyclic electron flow under drought stress by enhancing the expression of proteins related to photosystem I and cytochrome b6f. Shading significantly increased the expression of proteins related to the light harvesting complex, even under drought stress. In addition, shading increased the expression of proteins related to the oxygen evolution complex, plastocyanin, and ferredoxin–NADP+ reductase (FNR), thereby enhancing the efficiency of photosynthetic electron utilization. The shading and drought combination treatment appeared to enhance ginger's drought tolerance by reducing the expression of FNR and enhancing cyclic electron flow. Photosynthetic and fluorescence parameters showed that drought stress caused non-stomatal limitation of photosynthesis in ginger leaves. Drought stress also significantly reduced the quantum efficiency of photosystem II (Fv/Fm), the non-cyclic electron transfer efficiency of photosystem II (ϕPSII), and photochemical quenching (qP), while simultaneously increasing nonphotochemical quenching (NPQ). The addition of shading improved photosynthetic efficiency under drought. These results provide important baseline information on the photosynthetic mechanisms by which ginger responds to drought and shading. In addition, they provide a theoretical basis for the study of shade cultivation during the arid season.

Candidate Genes for Freezing and Drought Tolerance Selected on the Basis of Proteome Analysis in Doubled Haploid Lines of Barley.

Plant tolerance to environmental stress is determined by a very complicated network composed of many intra- and extracellular factors. The aim of this study was to select candidate genes involved in responses to freezing and drought in barley on the basis of previous proteomic studies and to analyze changes in their expression caused by application of both stress factors. Six candidate genes for freezing tolerance (namely the genes encoding elongation factor 1 alpha (EF1A), ferredoxin-NADP reductase, a 14-3-3a protein, β-fructofuranosidase, CBF2A and CBF4B) and six for drought tolerance (encoding transketolase, periplasmic serine protease, triosephosphate isomerase, a protein with a co-chaperon region (GroEs), pfam14200 and actin) were chosen arbitrarily on the basis of in silico bioinformatic analyses. The expression levels of these genes were measured under control and stress conditions in six DH (doubled haploid) lines with differing freezing and drought tolerance. The results of gene expression analysis confirmed the roles of the candidate genes preselected in this study on the basis of previous proteome analysis in contributing to the differences in freezing and drought tolerance observed in the studied population of DH lines of winter barley.

Transcriptomic and proteomic profiles of II YOU 838 (Oryza sativa) provide insights into heat stress tolerance in hybrid rice

Heat stress is an increasing threat to rice production worldwide. To investigate the mechanisms of heat tolerance in hybrid rice and their contributions to rice heterosis, we compared the transcriptome of the hybrid rice II YOU 838 (II8) with the transcriptomes of its parents Fu Hui 838 (F8) and II-32A (II3) after heat stress at 42 °C for 0 h, 24 h, 72 h and 120 h. We also performed a proteomic analysis in II8 after heat stress at 42 °C for 24 h. The transcriptome data revealed time-dependent gene expression patterns under the heat stress conditions, and the heat stress response of II8 was greatly different from those of its parents. Gene ontology analysis of the differentially expressed genes that were clustered using k-means clustering showed that most of the up-regulated genes were involved in responses to stimuli, cell communication, and metabolic and transcription factor activities, whereas the down-regulated genes were enriched in photosynthesis and signal transduction. Moreover, 35 unique differentially abundant proteins, including a basic helix-loop-helix transcription factor (bHLH96), calmodulin-binding transcription activator, heat shock protein (Hsp70), and chaperonin 60 (CPN60), were detected in the proteomic analysis of II8 under heat stress. The co-regulatory analysis revealed novel genes and pathways involved in heat tolerance, namely, ferredoxin-NADP reductase, peroxidases, mitogen-activated protein kinase kinase kinase, and heat shock factor (HSF)–Hsp network. Members of the Hsp and HSF families had over-dominant expression patterns in the hybrid compared with its parents, to help maintain the higher photosynthesis and antioxidant defense systems in the hybrid. Our study suggests that the complex HSF–Hsp regulatory network contribute to the heat tolerance of the hybrid rice.

Interaction of FNR with the Cytochrome b6f Complex: Thermodynamic Parameters

The cytochrome b6f complex, for which a crystal structure has been obtained at 2.5 Å resolution (Hasan and Cramer, Structure, 2014) has a central role in the regulation of oxygenic photosynthetic electron transport. The photosytem I peripheral electron transfer proteins ferredoxin (Fd) and ferredoxin NADP+ reductase (FNR) participate in the pathway of NADP+ reduction. Cyclic electron transfer pathway provides an alternate source of electron transfer-mediated membrane energization for ATP production. The present study focuses on the physical interaction of FNR with the cytochrome b6f complex, with relevance to the participation of FNR in a cyclic pathway. The presence of FNR in purified cytochrome b6f complex has been documented and found to be present in sub-stoichiometric levels (Zhang et al. JBC, 2001). It is known that linear electron transfer does not require direct interaction of the cytochrome complex with FNR. In the present study, isothermal calorimetry (ITC) was used to detect and characterize the interaction of FNR with the dimeric cytochrome b6f complex, derived from a plant (spinach) source, in detergent solution. The interaction is relatively weak, with thermodynamic parameters: Kd, 48 μM; ΔH, −23.3 kJ/mol; ΔS, 3.3 J/mol K, at pH 6.5, and ionic strength of 170 mM. It is of interest that such interactions were not detected at pH 8.0, implying a significant pH dependence. Far-UV circular dichroism (CD) analysis of the structure stability of FNR was employed to analyze the pH dependence of the interactions and the effect of detergent. This analysis reveals the decrease in structure stability of FNR and less efficient interaction with the b6f complex in the presence of the detergent UDM. Study supported by U.S. Department of Energy (DOE DE-SC0018238).

A Double-Activity (Green Algae Toxicity and Bacterial Genotoxicity) 3D-QSAR Model Based on the Comprehensive Index Method and Its Application in Fluoroquinolones' Modification.

The comparative molecular similarity index analysis (CoMSIA) model of double-activity quinolones targeting green algae toxicity and bacterial genotoxicity (8:2) was constructed in this paper on the basis of the comprehensive index method. The contour maps of the model were analyzed for molecular modifications with high toxicities. In the CoMSIA model, the optimum number of components n was 7, the cross-validated q2 value was 0.58 (>0.5), the standard deviation standard error of estimate (SEE) was 0.02 (<0.95), F was 1265.33, and the non-cross-validated R2 value was 1 (>0.9), indicating that the model had a good fit and predicting ability. The scrambling stability test parameters Q2, cross-validated standard error of prediction (cSDEP), and dq2/dr2yy were 0.54, 0.25, and 0.8 (<1.2), respectively, indicating that the model had good stability. The external verification coefficient r2pred was 0.73 (>0.6), and standard error of prediction (SEP) was 0.17, indicating that the model had a good external prediction ability. The contribution rates of the steric fields, electrostatic fields, hydrophobic fields, hydrogen bond donor, and acceptor fields were 10.9%, 19.8%, 32.7%, 13.8%, and 22.8%, respectively. Large volume groups were selected for modification of ciprofloxacin (CIP), and the derivatives with increased double-activity characterization values were screened; the increase ratio ranged from 12.31–19.09%. The frequency of derivatives were positive and total energy, bioaccumulation, and environmental persistence was reduced, indicating that the CIP derivatives had good environmental stability and friendliness. Predicted values and CoMSIA model constructed of single activities showed that the CoMSIA model of double activities had accuracy and reliability. In addition, the total scores of the derivatives docking with the D1 protein, ferredoxin-NADP (H) reductases (FNRs), and DNA gyrase increased, indicating that derivatives can be toxic to green algae by affecting the photosynthesis of green algae. The mechanism behind the bactericidal effect was also explained from a molecular perspective.

Interprotein electron transfer biohybrid system for photocatalytic H2 production.

Worldwide there is a large research investment in developing solar fuel systems as clean and sustainable sources of energy. The fundamental mechanisms of natural photosynthesis can provide a source of inspiration for these studies. Photosynthetic reaction center (RC) proteins capture and convert light energy into chemical energy that is ultimately used to drive oxygenic water-splitting and carbon fixation. For the light energy to be used, the RC communicates with other donor/acceptor components via a sophisticated electron transfer scheme that includes electron transfer reactions between soluble and membrane bound proteins. Herein, we reengineer an inherent interprotein electron transfer pathway in a natural photosynthetic system to make it photocatalytic for aqueous H2 production. The native electron shuttle protein ferredoxin (Fd) is used as a scaffold for binding of a ruthenium photosensitizer and H2 catalytic function is imparted to its partner protein, ferredoxin-NADP+-reductase (FNR), by attachment of cobaloxime molecules. We find that this 2-protein biohybrid system produces H2 in aqueous solutions via light-induced interprotein electron transfer reactions (TON > 2500 H2/FNR), providing insight about using native protein-protein interactions as a method for fuel generation.

Proteomic Analysis of the Protective Effect of Sodium Nitroprusside on Leaves of Barley Stressed by Salinity

Objective: The salinization of agricultural soils poses a serious challenge across the world. Although recent studies have shown that exogenous sodium nitroprusside (SNP) application can alleviate the harmful effects of salinity, the roles of SNP in the regulation of proteomic changes remain poorly understood. Materials and Methods: To unravel the protective roles of exogenous SNP in alleviating salt-induced damage in barley (Hordeum vulgare L.), proteomic analysis was carried out on the leaves of seedlings exposed to 100 mM NaCl stress following 200 µM SNP pre-treatment. Results: Our results indicated that SNP pre-treatment restored the seedling growth reduced by salinity stress. Comparing 2-DE gels from the treatments showed that 24 proteins were differentially accumulated under SNP and/or NaCl stress treatments. Among them, 15 proteins were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Gene ontology analysis demonstrated that several pathways were regulated by SNP and/or NaCl treatments, including photosynthesis, protein metabolism, stress defense, and energy metabolism. Exogenous SNP increased the expression levels of 20 kDa chaperonin, proteasome subunit beta type-2, 2-Cys peroxiredoxin BAS1, ferredoxin-NADP reductase, thiazole biosynthetic enzyme 1-1, S-adenosylmethionine synthetase 3, and elongation factor Tu proteins in the leaves of barley seedlings under NaCl stress. Conclusion: Our results indicate that SNP pre-treatment may induce salinity tolerance through regulation of photosynthesis, activation of stress defence, degradation of damaged proteins, and the promoting of the synthesis of polyamines, proline, and GABA.

NADP(H) allosterically regulates the interaction between ferredoxin and ferredoxin-NADP+ reductase.

Ferredoxin‐NADP+ reductase (FNR) in plants receives electrons from ferredoxin (Fd) at the end of the photosynthetic electron transfer chain and converts NADP+ to NADPH. The interaction between Fd and FNR in plants was previously shown to be attenuated by NADP(H). Here, we investigated the molecular mechanism of this phenomenon using maize FNR and Fd, as the three‐dimensional structure of this complex is available. NADPH, NADP+, and 2′5′‐ADP differentially affected the interaction, as revealed through kinetic and physical binding analyses. Site‐directed mutations of FNR which change the affinity for NADPH altered the affinity for Fd in the opposite direction to that for NADPH. We propose that the binding of NADP(H) causes a conformational change of FNR which is transferred to the Fd‐binding region through different domains of FNR, resulting in allosteric changes in the affinity for Fd.