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Metabolic engineering is a critical biotechnological approach in addressing global energy and environment challenges. Most engineering efforts, however, consist of laborious and inefficient trial-and-error of target pathways, due in part to the lack of methodologies that can comprehensively assess pathway properties in thermodynamics and kinetics. Metabolic engineering can benefit from computational tools that evaluate feasibility, expense and stability of non-natural metabolic pathways. Such tools can also help us understand natural pathways and their regulation at systems level. Here we introduce a computational toolbox, PathParser, which, for the first time, integrates multiple important functions for pathway analysis including thermodynamics analysis, kinetics-based protein cost optimization and robustness analysis. Specifically, PathParser enables optimization of the driving force of a pathway by minimizing the Gibbs free energy of least thermodynamically favorable reaction. In addition, based on reaction thermodynamics and enzyme kinetics, it can compute the minimal enzyme protein cost that supports metabolic flux, and evaluate pathway stability and flux in response to enzyme concentration perturbations. In a demo analysis of the Calvin-Benson-Bassham cycle and photorespiration pathway in the model cyanobacterium Synechocystis PCC 6803, the computation results are corroborated by experimental proteomics data as well as metabolic engineering outcomes. This toolbox may have broad application in metabolic engineering and systems biology in other microbial systems., (Copyright © 2019 International Metabolic Engineering Society. All rights reserved.)

Iron intoxications induce severe oxidative stress by producing reactive oxygen species (ROS) in cyanobacteria, leading to membrane lipid peroxidation, altered morphology, impaired photosynthesis and other oxidative stress injuries. Given these stresses, mitigation of ROS is a prerequisite for all aerobic organisms. Study of siderophilic cyanobacterium Leptolyngbya strain JSC-1 inhabiting iron-rich hot springs may provide insight into the mechanism of iron homeostasis and alleviation of oxidative stress. In this study, we investigated the morphophysiological and molecular mechanisms enabling this cyanobacterium to cope with iron-induced oxidative stress. Strain JSC-1 biomineralized extracellular iron via an exopolymeric sheath (acting as a first line of defense) and intracellular iron via polyphosphate inclusions (second line of defense), thus minimizing the burden of free ferric ions. Physiological parameters, SOD, CAT and POD activities, bacterioferritin and total protein contents fluctuated in response to iron elevation, displaying a third line of defense to mitigate ROS. Differential gene expression analysis of JSC-1 indicated up-regulation of 94 and 125 genes and down-regulation of 89 and 183 genes at low (4 μM) and high (400 μM) iron concentration, respectively. The differentially expressed genes (DEGs) were enriched in 100 KEGG pathways and were found to be involved in lipopolysaccharide and fatty acid biosynthesis, starch, sucrose, chlorophyll and other metabolic pathways. Together with metabolic reprogramming (fourth line of defense), JSC-1 established a unique multiline defense system that allows JSC-1 to withstand severe oxidative stress. These findings also provide insight into potential survival strategies of ancient microorganisms inhabiting similar environment present in early earth history., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Although some elements of the photosynthetic light reactions might appear to be ideal, the overall efficiency of light conversion to biomass has not been optimized during evolution. Because crop plants are depleted of genetic diversity for photosynthesis, efforts to enhance its efficiency with respect to light conversion to yield must generate new variation. In principle, three sources of natural variation are available: (i) rare diversity within extant higher plant species, (ii) photosynthetic variants from algae, and (iii) reconstruction of no longer extant types of plant photosynthesis. Here, we argue for a novel approach that outsources crop photosynthesis to a cyanobacterium that is amenable to adaptive evolution. This system offers numerous advantages, including a short generation time, virtually unlimited population sizes and high mutation rates, together with a versatile toolbox for genetic manipulation. On such a synthetic bacterial platform, 10 000 years of (crop) plant evolution can be recapitulated within weeks. Limitations of this system arise from its unicellular nature, which cannot reproduce all aspects of crop photosynthesis. But successful establishment of such a bacterial host for crop photosynthesis promises not only to enhance the performance of eukaryotic photosynthesis but will also reveal novel facets of the molecular basis of photosynthetic flexibility.This article is part of the themed issue 'Enhancing photosynthesis in crop plants: targets for improvement'., (© 2017 The Author(s).)

Florfenicol (FF) and thiamphenicol (TAP) are two typical pharmaceuticals used widely as therapeutica antibiotic agents in aquaculture. However, little is known about the potential adverse effects of these two antibiotics on non-target organisms in the aquatic ecosystem. In this study we investigated the effects of FF and TAP on photosynthesis and the antioxidant system of the cyanobacteria Microcystis flos-aquae. Over a concentration range of 0.001-1μg/L, the results showed that both FF and TAP significantly increased the chlorophyll a content of M. flos-aquae, while the superoxide dismutase (SOD) activity, catalase (CAT) activity and the levels of malondialdehyde (MDA) changed slightly. In contrast, the chlorophyll a content of M. flos-aqua was significantly inhibited (p<0.01) at high concentrations (>1μg/L) of FF and TAP, reaching a 46% inhibition level at 50μg/L FF and 56% inhibition at 100μg/L TAP. At the same time, the activities of SOD and CAT along with MDA content also increased significantly (p<0.01), indicating that the high concentrations of both FF and TAP led to oxidative stress in the algae. In addition, the M. flos-aquae fluorescence parameters (Fv/Fm, Fv/Fo, alpha, ETRmax and Ik) increased with increasing concentration of both FF and TAP, which may be the result of the increasing photoprotection capacity., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Cyanobacteria are the oldest known life form inhabiting Earth and the only prokaryotes capable of performing oxygenic photosynthesis. Synechocystis sp. PCC 6803 (Synechocystis) is a model cyanobacterium used extensively in research on photosynthesis and environmental adaptation. Posttranslational protein modification by lysine acetylation plays a critical regulatory role in both eukaryotes and prokaryotes; however, its extent and function in cyanobacteria remain unexplored. Herein, we performed a global acetylome analysis on Synechocystis through peptide prefractionation, antibody enrichment, and high accuracy LC-MS/MS analysis; identified 776 acetylation sites on 513 acetylated proteins; and functionally categorized them into an interaction map showing their involvement in various biological processes. Consistent with previous reports, a large fraction of the acetylation sites are present on proteins involved in cellular metabolism. Interestingly, for the first time, many proteins involved in photosynthesis, including the subunits of phycocyanin (CpcA, CpcB, CpcC, and CpcG) and allophycocyanin (ApcA, ApcB, ApcD, ApcE, and ApcF), were found to be lysine acetylated, suggesting that lysine acetylation may play regulatory roles in the photosynthesis process. Six identified acetylated proteins associated with photosynthesis and carbon metabolism were further validated by immunoprecipitation and Western blotting. Our data provide the first global survey of lysine acetylation in cyanobacteria and reveal previously unappreciated roles of lysine acetylation in the regulation of photosynthesis. The provided data set may serve as an important resource for the functional analysis of lysine acetylation in cyanobacteria and facilitate the elucidation of the entire metabolic networks and photosynthesis process in this model cyanobacterium.

4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR), also known as isoprenoid synthesis H (IspH) or lysis-tolerant B (LytB), catalyzes the last step of the methylerythritol phosphate pathway to synthesize isopentenyl diphosphate and dimethylallyl diphosphate. The structure and reaction mechanism of IspH have been actively investigated in Escherichia coli but little is known in plants. Compared with the bacterial IspH, cyanobacterial and plant HDRs all contain an extra N-terminal conserved domain (NCD) that is essential for their function. Tyr72 in the NCD and several plant-specific residues around the central active site are critical for Arabidopsis HDR function. These results suggest that the structure and reaction mechanism of HDR/IspH may be different between plants and bacteria. The E. coli IspH is an iron-sulfur protein that is sensitive to oxygen. It is possible that the cyanobacterial HDR may independently evolve from the common ancestor of prokaryotes to obtain the NCD, which may protect the enzyme from high concentration of oxygen during photosynthesis.

A phycocyanin-deletion mutant of Synechocystis (cyanobacteria) was generated upon replacement of the CPC-operon with a kanamycin resistance cassette. The Δcpc transformant strains (Δcpc) exhibited a green phenotype, compared to the blue-green of the wild type (WT), lacked the distinct phycocyanin absorbance at 625nm, and had a lower Chl per cell content and a lower PSI/PSII reaction center ratio compared to the WT. Molecular and genetic analyses showed replacement of all WT copies of the Synechocystis DNA with the transgenic version, thereby achieving genomic DNA homoplasmy. Biochemical analyses showed the absence of the phycocyanin α- and β-subunits, and the overexpression of the kanamycin resistance NPTI protein in the Δcpc. Physiological analyses revealed a higher, by a factor of about 2, intensity for the saturation of photosynthesis in the Δcpc compared to the WT. Under limiting intensities of illumination, growth of the Δcpc was slower than that of the WT. This difference in the rate of cell duplication diminished gradually as growth irradiance increased. Identical rates of cell duplication of about 13h for both WT and Δcpc were observed at about 800μmolphotonsm(-2)s(-1) or greater. Culture productivity analyses under simulated bright sunlight and high cell-density conditions showed that biomass accumulation by the Δcpc was 1.57-times greater than that achieved by the WT. Thus, the work provides first-time direct evidence of the applicability of the Truncated Light-harvesting Antenna (TLA)-concept in cyanobacteria, entailing substantial improvements in the photosynthetic efficiency and productivity of mass cultures upon minimizing the phycobilisome light-harvesting antenna size., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Ultraviolet (UV) A radiation (315–400 nm) is the predominant component of solar UV radiation that reaches the Earth’s surface. However, the underlying mechanisms of the positive effects of UV-A on photosynthetic organisms have not yet been elucidated. In this study, we investigated the effects of UV-A radiation on the growth, photosynthetic ability, and metabolome of the edible cyanobacterium Nostoc sphaeroides. Exposures to 5–15 W m−2 (15–46 µmol photons m−2 s−1) UV-A and 4.35 W m−2 (20 μmol photons m−2 s−1) visible light for 16 days significantly increased the growth rate and biomass production of N. sphaeroides cells by 18%–30% and 15%–56%, respectively, compared to the non-UV-A-acclimated cells. Additionally, the UV-A-acclimated cells exhibited a 1.8-fold increase in the cellular nicotinamide adenine dinucleotide phosphate (NADP) pool with an increase in photosynthetic capacity (58%), photosynthetic efficiency (24%), QA re-oxidation, photosystem I abundance, and cyclic electron flow (87%), which further led to an increase in light-induced NADPH generation (31%) and ATP content (83%). Moreover, the UV-A-acclimated cells showed a 2.3-fold increase in ribulose-1,5-bisphosphate carboxylase/oxygenase activity, indicating an increase in their carbon-fixing capacity. Gas chromatography–mass spectrometry-based metabolomics further revealed that UV-A radiation upregulated the energy-storing carbon metabolism, as evidenced by the enhanced accumulation of sugars, fatty acids, and citrate in the UV-A-acclimated cells. Therefore, our results demonstrate that UV-A radiation enhances energy flow and carbon assimilation in the cyanobacterium N. sphaeroides. IMPORTANCE Ultraviolet (UV) radiation exerts harmful effects on photo-autotrophs; however, several studies demonstrated the positive effects of UV radiation, especially UV-A radiation (315–400 nm), on primary productivity. Therefore, understanding the underlying mechanisms associated with the promotive effects of UV-A radiation on primary productivity can facilitate the application of UV-A for CO2 sequestration and lead to the advancement of photobiological sciences. In this study, we used the cyanobacterium Nostoc sphaeroides, which has an over 1,700-year history of human use as food and medicine, to explore its photosynthetic acclimation response to UV-A radiation. As per our knowledge, this is the first study to demonstrate that UV-A radiation increases the biomass yield of N. sphaeroides by enhancing energy flow and carbon assimilation. Our findings provide novel insights into UV-A-mediated photosynthetic acclimation and provide a scientific basis for the application of UV-A radiation for optimizing light absorption capacity and enhancing CO2 sequestration in the frame of a future CO2 neutral, circular, and sustainable bioeconomy. [ABSTRACT FROM AUTHOR]

Carbon acquisition, assimilation and storage in eukaryotic microalgae and cyanobacteria occur in multiple compartments that have been characterised by the location of the enzymes involved in these functions. These compartments can be delimited by bilayer membranes, such as the chloroplast, the lumen, the peroxisome, the mitochondria or monolayer membranes, such as lipid droplets or plastoglobules. They can also originate from liquid–liquid phase separation such as the pyrenoid. Multiple exchanges exist between the intracellular microcompartments, and these are reviewed for the CO2 concentration mechanism, the Calvin–Benson–Bassham cycle, the lipid metabolism and the cellular energetic balance. Progress in microscopy and spectroscopic methods opens new perspectives to characterise the molecular consequences of the location of the proteins involved, including intrinsically disordered proteins. [ABSTRACT FROM AUTHOR]

Synechocystis sp. PCC 6803 (hereafter: Synechocystis) is a model organism for studying photosynthesis, energy metabolism, and environmental stress. Although known as the first fully sequenced phototrophic organism, Synechocystis still has almost half of its proteome without functional annotations. In this study, by using co-fractionation coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS), we define 291 multi-protein complexes, encompassing 24,092 protein–protein interactions (PPIs) among 2062 distinct gene products. This information not only reveals the roles of photosynthesis in metabolism, cell motility, DNA repair, cell division, and other physiological processes, but also shows how protein functions vary from bacteria to higher plants due to changes in interaction partners. It also allows us to uncover the functions of hypothetical proteins, such as Sll0445, Sll0446, and Sll0447 involved in photosynthesis and cell motility, and Sll1334 involved in regulation of fatty acid biogenesis. Here we present the most extensive PPI data for Synechocystis so far, which provide critical insights into fundamental molecular mechanisms in cyanobacteria.

Summary: Cytochrome (Cyt) b559 is a key component of the photosystem II complex (PSII) that is essential for its proper functioning and assembly. Site‐directed mutants of the model cyanobacterium Synechocystis sp. PCC6803 with mutated heme axial ligands of Cyt b559 have little PSII and are therefore unable to grow photoautotrophically.Here we describe two types of Synechocystis autotrophic transformants that retained the same mutations in Cyt b559 but are able to accumulate PSII and grow photoautotrophically. Whole‐genome sequencing revealed that all of these autotrophic transformants carried a variable number of tandem repeats (from 5 to 15) of chromosomal segments containing the psbEFLJ operon.RNA‐seq analysis showed greatly increased transcript levels of the psbEFLJ operon in these autotrophic transformants. Multiple copies of the psbEFLJ operon in these transformants were only maintained during autotrophic growth, while its copy numbers gradually decreased under photoheterotrophic conditions. Two‐dimensional PAGE analysis of membrane proteins revealed a strong deficiency in PSII complexes in the Cyt b559 mutants that was reversed in the autotrophic transformants.These results illustrate how tandem gene amplification restores PSII accumulation and photoautotrophic growth in Cyt b559 mutants of cyanobacteria, and may serve as an important adaptive mechanism for cyanobacterial survival. [ABSTRACT FROM AUTHOR]

Abstract Background Cyanobacteria are ideal model organisms to exploit photosynthetically derived electrons or fixed carbon for the biotechnological synthesis of high value compounds and energy carriers. Much effort is spent on the rational design of heterologous pathways to produce value-added chemicals. Much less focus is drawn on the basic physiological responses and potentials of phototrophs to deal with natural or artificial electron and carbon sinks. However, an understanding of how electron sinks influence or regulate cellular physiology is essential for the efficient application of phototrophic organisms in an industrial setting, i.e., to achieve high productivities and product yields. Results The physiological responses of the cyanobacterium Synechocystis sp. PCC 6803 to electron sink variation were investigated in a systematic and quantitative manner. A variation in electron demand was achieved by providing two N sources with different degrees of reduction. By additionally varying light and CO2 availabilities, steady state conditions with strongly differing source–sink ratios were established. Balancing absorbed photons and electrons used for different metabolic processes revealed physiological responses to sink/source ratio variation. Surprisingly, an additional electron sink under light and thus energy limitation was found not to hamper growth, but was compensated by improved photosynthetic efficiency and activity. In the absence of carbon and light limitation, an increase in electron demand even stimulated carbon assimilation and growth. Conclusion The metabolism of Synechocystis sp. PCC 6803 is highly flexible regarding the compensation of additional electron demands. Under light limitation, photosynthesis obviously does not necessarily run at its maximal capacity, possibly for the sake of robustness. Increased electron demands can even boost photosynthetic activity and growth.

Soil salinity in nature is generally mixed type; however, most of the studies on salt toxicity are performed with NaCl and little is known about sulfur type of salinity (Na2SO4). Present study discerns the physiologic mechanisms responsible for salt tolerance in salt-adapted Anabaena fertilissima, and responses of directly stressed parent cells to NaCl and NaCl+Na2SO4 mixture. NaCl at 500 mM was lethal to the cyanobacterium, whereas salt-adapted cells grew luxuriantly. Salinity impaired gross photosynthesis, electron transport activities, and respiration in parent cells, but not in the salt-adapted cells, except a marginal increase in PSI activity. Despite higher Na+ concentration in the salt mixture, equimolar NaCl appeared more inhibitive to growth. Sucrose and trehalose content and antioxidant activities were maximal in 250 mM NaCl-treated cells, followed by salt mixture and was almost identical in salt-adapted (exposed to 500 mm NaCl) and control cells, except a marginal increase in ascorbate peroxidase activity and an additional fourth superoxide dismutase isoform. Catalase isoform of 63 kDa was induced only in salt-stressed cells. Salinity increased the uptake of intracellular Na+ and Ca2+ and leakage of K+ in parent cells, while cation level in salt-adapted cells was comparable to control. Though there was differential increase in intracellular Ca2+ under different salt treatments, ratio of Ca2+/Na+ remained the same. It is inferred that stepwise increment in the salt concentration enabled the cyanobacterium to undergo priming effect and acquire robust and efficient defense system involving the least energy. [ABSTRACT FROM AUTHOR]

Eukaryotes co-opted photosynthetic carbon fixation from prokaryotes by engulfing a cyanobacterium and stably integrating it as a photosynthetic organelle (plastid) in a process known as primary endosymbiosis. The sheer complexity of interactions between a plastid and the surrounding cell that started to evolve over 1 billion years ago, make it challenging to reconstruct intermediate steps in organelle evolution by studying extant plastids. Recently, the photosynthetic amoeba Paulinella chromatophora was identified as a much sought-after intermediate stage in the evolution of a photosynthetic organelle. This article reviews the current knowledge on this unique organism. In particular it describes how the interplay of reductive genome evolution, gene transfers, and trafficking of host-encoded proteins into the cyanobacterial endosymbiont contributed to transform the symbiont into a nascent photosynthetic organelle. Together with recent results from various other endosymbiotic associations a picture emerges that lets the targeting of host-encoded proteins into bacterial endosymbionts appear as an early step in the establishment of an endosymbiotic relationship that enables the host to gain control over the endosymbiont.

Biological production of silver nanoparticles (SNP) has recently received considerable attention because of their therapeutic applications. The present study reports the production and characterization of spherical-thermostable SNP from Spirulina platensis. Wet biomass harvested from optimized logarithmic-phase culture was used for production of SNP in 1, 3, or 5 mM silver nitrate solution. Formation and concentration of spherical SNP was highest when S. platensis was treated with 3 mM silver nitrate. Fourier transform infrared spectra of SNP indicate that vanillin, coumarins, tannins, amide, and glycogen may act as stabilizing agents for bioreduction. The crystalline nature of the produced SNP was evidenced from X-ray diffractometer analysis. Weight loss of SNP occurred at 210°C, 310°C, and 510°C as shown by thermogravimetric analysis. This study shows that S. platensis may be used as an efficient tool for production of spherical, crystalline, and thermostable SNP. [ABSTRACT FROM AUTHOR]

Nostoc calcicola, a halotolerant alkaliphilic, nitrogen-fixing cyanobacterium, was grown under various NaCl concentrations (from 500 to 2000 mM) at alkaline pH. In cyanobacteria, physiological and biochemical responses were correlated with the external salt stress. The optimal condition for cell growth of N. calcicola isolated from alkaline 'Usar' soils of northern India was 500 mM NaCl at pH 10.5, accompanied with an increase in photosynthetic O evolution, carbohydrate content and activities of nitrate reductase, glutamine synthetase, photosystem I and photosystem II. Under salt stress conditions above 500 mM NaCl, cell growth and carbohydrate contents reduced. The results indicate that production of carbohydrates under salinity stress at alkaline pH acts as major mechanism of salt tolerance in cyanobacteria. [ABSTRACT FROM AUTHOR]

Cyanobacteria are an important group of photoautotrophic bacteria that have been engineered and used to produce a wide range of biomaterials and biofuels, which are usually derived from important intermediates of the central metabolic pathways. In this study, the production of poly-3-hydroxybutyrate and poly-3-hydroxybutyrate-co-4-hydroxybutyrate in cyanobacteria was studied, and metabolic engineering strategies to improve the yields were also investigated. The genes involved in the biosynthetic pathway for poly-3-hydroxybutyrate from Chlorogloeopsis fritschii PCC 9212 were introduced into Synechococcus sp. PCC 7002, and the resulting strain was able to accumulate 2.77% of total cell dry weight as poly-3-hydroxybutyrate. When the ccmR gene was deleted in this strain, the yield of poly-3-hydroxybutyrate increased to 3.97% of total cell dry weight. A biosynthetic pathway for the production of 4-hydroxybutyryl-CoA was also constructed and introduced into the poly-3-hydroxybutyrate-producing strain. The resulting strain was able to produce ~4.5% of total cell dry weight as poly-3-hydroxybutyrate-co-4-hydroxybutyrate, in which 4-hydroxybutyrate accounted for ~12% of the co-polymer. These results demonstrate that poly-3-hydroxybutyrate-co-4-hydroxybutyrate can be produced in cyanobacteria and confirm that succinic semialdehyde is an important TCA cycle metabolite in cyanobacteria. This study also demonstrates the potential for future metabolic engineering in cyanobacteria that is based on recently discovered metabolites. [ABSTRACT FROM AUTHOR]

Eukaryotes co-opted photosynthetic carbon fixation from prokaryotes by engulfing a cyanobacterium and stably integrating it as a photosynthetic organelle (plastid) in a process known as primary endosymbiosis. The sheer complexity of interactions between a plastid and the surrounding cell that started to evolve over 1 billion years ago, make it challenging to reconstruct intermediate steps in organelle evolution by studying extant plastids. Recently, the photosynthetic amoeba Paulinella chromatophora was identified as a much sought-after intermediate stage in the evolution of a photosynthetic organelle. This article reviews the current knowledge on this unique organism. In particular it describes how the interplay of reductive genome evolution, gene transfers, and trafficking of host-encoded proteins into the cyanobacterial endosymbiont contributed to transform the symbiont into a nascent photosynthetic organelle. Together with recent results from various other endosymbiotic associations a picture emerges that lets the targeting of host-encoded proteins into bacterial endosymbionts appear as an early step in the establishment of an endosymbiotic relationship that enables the host to gain control over the endosymbiont. [ABSTRACT FROM AUTHOR]

Highlights: [•] Leptolyngbya foveolarum tolerated cartap up to 80ppm. [•] Low dose (20ppm) of cartap stimulated growth, photosynthetic pigments, photosynthesis and respiration. [•] Higher dose (40 and 60ppm) of insecticide inhibited same parameters. [•] Nitrogen assimilation was affected in dose dependent manner by cartap. [•] The organism lyzed in nitrogen depleted medium but survived when cartap was supplemented. [Copyright &y& Elsevier]

Plastocyanin, encoded by the petE gene, can transfer electrons to photosystem I ( PSI) and cytochrome c oxidase during photosynthetic and respiratory metabolism in cyanobacteria. We constructed a petE mutant of Synechocystis sp. strain PCC 6803 and investigated its phenotypic properties under different light conditions. When cultured under continuous light, inactivation of petE accelerated the plastoquinone pool reoxidation, slowed the reoxidation rate of the primary quinone-type acceptor, and decreased the connectivity factor between the individual photosystem II ( PSII) photosynthetic units. Compared with the wild-type control, the petE mutant showed a decrease in its PSI/ PSII fluorescence ratio and an increase in its dark respiration rate. When cultured under a light-dark photoperiod, the petE mutation caused an increase in the phycocyanin to chlorophyll ratio. Consequently, the mutant line was a darker blue than its wild-type counterpart. Moreover, the petE mutation increased the efficiency of light capture, nonphotochemical quenching, and linear electron transport activity, but decreased the functional absorption cross section of PSII. These results suggest that plastocyanin is involved in regulating the redox state of the photosynthetic electron transfer chain, and the petE mutation can induce interesting phenotypic properties that are specific to the light-dark photoperiod. [ABSTRACT FROM AUTHOR]

Abstract: The individual and combined toxicities of amoxicillin, erythromycin, levofloxacin, norfloxacin and tetracycline have been examined in two organisms representative of the aquatic environment, the cyanobacterium Anabaena CPB4337 as a target organism and the green alga Pseudokirchneriella subcapitata as a non-target organism. The cyanobacterium was more sensitive than the green alga to the toxic effect of antibiotics. Erythromycin was highly toxic for both organisms; tetracycline was more toxic to the green algae whereas the quinolones levofloxacin and norfloxacin were more toxic to the cyanobacterium than to the green alga. Amoxicillin also displayed toxicity to the cyanobacterium but showed no toxicity to the green alga. The toxicological interactions of antibiotics in the whole range of effect levels either in binary or multicomponent mixtures were analyzed using the Combination Index (CI) method. In most cases, synergism clearly predominated both for the green alga and the cyanobacterium. The CI method was compared with the classical models of additivity Concentration Addition (CA) and Independent Action (IA) finding that CI could accurately predict deviations from additivity. Risk assessment was performed by calculating the ratio between Measured Environmental Concentration (MEC) and the Predicted No Effect Concentration (PNEC). A MEC/PNEC ratio higher than 1 was found for the binary erythromycin and tetracycline mixture in wastewater effluents, a combination which showed a strong synergism at low effect levels in both organisms. From the tested antibiotic mixtures, it can be concluded that certain specific combinations may pose a potential ecological risk for aquatic ecosystems with the present environmentally measured concentrations. [Copyright &y& Elsevier]

Abstract: X-ray crystal structures of the isolated phycobiliprotein components of the phycobilisome have provided high resolution details to the description of this light harvesting complex at different levels of complexity and detail. The linker-independent assembly of trimers into hexamers in crystal lattices of previously determined structures has been observed in almost all of the phycocyanin (PC) and allophycocyanin (APC) structures available in the Protein Data Bank. In this paper we describe the X-ray crystal structures of PC and APC from Synechococcus elongatus sp. PCC 7942, PC from Synechocystis sp. PCC 6803 and PC from Thermosynechococcus vulcanus crystallized in the presence of urea. All five structures are highly similar to other PC and APC structures on the levels of subunits, monomers and trimers. The Synechococcus APC forms a unique loose hexamer that may show the structural requirements for core assembly and rod attachment. While the Synechococcus PC assembles into the canonical hexamer, it does not further assemble into rods. Unlike most PC structures, the Synechocystis PC fails to form hexamers. Addition of low concentrations of urea to T. vulcanus PC inhibits this proteins propensity to form hexamers, resulting in a crystal lattice composed of trimers. The molecular source of these differences in assembly and their relevance to the phycobilisome structure is discussed. [Copyright &y& Elsevier]

Glutathione biosynthesis catalysed by GCL (glutamate-cysteine ligase) and GS (glutathione synthetase) is essential for maintaining redox homoeostasis and protection against oxidative damage in diverse eukaroytes and bacteria. This biosynthetic pathway probably evolved in cyanobacteria with the advent of oxygenic photosynthesis, but the biochemical characteristics of progenitor GCLs and GSs in these organisms are largely unexplored. In the present study we examined SynGCL and SynGS from Synechocystis sp. PCC 6803 using steady-state kinetics. Although SynGCL shares ~15% sequence identity with the enzyme from plants and α-proteobacteria, sequence comparison suggests that these enzymes share similar active site residues. Biochemically, SynGCL lacks the redox regulation associated with the plant enzymes and functions as a monomeric protein, indicating that evolution of redox regulation occurred later in the green lineage. Site-directed mutagenesis of SynGCL establishes this enzyme as part of the plant-like GCL family and identifies a catalytically essential arginine residue, which is structurally conserved across all forms of GCLs, including those from non-plant eukaryotes and γ -proteobacteria. A reaction mechanism for the synthesis of γ -glutamylcysteine by GCLs is proposed. Biochemical and kinetic analysis of SynGS reveals that this enzyme shares properties with other prokaryotic GSs. Initial velocity and product inhibition studies used to examine the kinetic mechanism of SynGS suggest that it and other prokaryotic GSs uses a random ter-reactant mechanism for the synthesis of glutathione. The present study provides new insight on the molecular mechanisms and evolution of glutathione biosynthesis; a key process required for enhancing bioenergy production in photosynthetic organisms. [ABSTRACT FROM AUTHOR]

Abstract: Psb28 is an extrinsic protein of photosystem II (PSII), which is conserved among photosynthetic organisms from cyanobacteria to higher plants. A unicellular cyanobacterium, Synechocystis sp. PCC 6803, has two homologs of Psb28, Psb28-1 and Psb28-2. However, the role of these proteins remains poorly understood. In this study, we disrupted the psb28-1 (sll1398) and psb28-2 (slr1739) genes in wild-type Synechocystis sp. PCC 6803 and examined their photosynthetic properties to elucidate the physiological role of Psb28 in photosynthesis. We also disrupted the psb28-1 gene in a dgdA mutant defective in the biosynthesis of digalactosyldiacylglycerol, in which Psb28-1 significantly accumulates in PSII. The disruption of the psb28-1 gene in the wild-type resulted in growth retardation under high-light conditions at high temperatures with a low rate of restoration of photodamaged photosynthetic machinery. Similar phenomena were observed at normal growth temperatures in the psb28-1/dgdA double mutant. In contrast, disruption of psb28-2 in the wild-type and dgdA mutant did not affect host strain phenotype, suggesting that Psb28-2 does not contribute to the recovery of PSII. In addition, protein analysis using strains expressing His-tagged Psb28-1 revealed that Psb28-1 is mainly associated with the CP43-less PSII monomer. In the dgdA mutant, the CP43-less PSII monomer accumulated to a greater extent than in the wild-type, and its accumulation caused greater accumulation of Psb28-1 in PSII. These results demonstrate that Psb28-1 plays an important role in PSII repair through association with the CP43-less monomer, particularly at high temperatures. [Copyright &y& Elsevier]

A large variety of new energy-generating technologies are being developed in an effort to reduce global dependence on fossil fuels, and to reduce the carbon footprint of energy generation. The term 'biological photovoltaic system' encompasses a broad range of technologies which all employ biological material that can harness light energy to split water, and then transfer the resulting electrons to an anode for power generation or electrosynthesis. The use of whole cyanobacterial cells is a good compromise between the requirements of the biological material to be simply organized and transfer electrons efficiently to the anode, and also to be robust and able to self-assemble and self-repair. The principle that photosynthetic bacteria can generate and transfer electrons directly or indirectly to an anode has been demonstrated by a number of groups, although the power output obtained from these devices is too low for biological photovoltaic devices to be useful outside the laboratory. Understanding how photosynthetically generated electrons are transferred through and out of the organism is key to improving power output, and investigations on this aspect of the technology are the main focus of the present review. [ABSTRACT FROM AUTHOR]

Abstract: Cytochrome c550 (cyt c550) is a membrane component of the PSII complex in cyanobacteria and some eukaryotic algae, such as red and brown algae. Cyt c550 presents a bis-histidine heme coordination which is very unusual for monoheme c-type cytochromes. In PSII, the cyt c550 with the other extrinsic proteins stabilizes the binding of Cl− and Ca2+ ions to the oxygen evolving complex and protects the Mn4Ca cluster from attack by bulk reductants. The role (if there is one) of the heme of the cyt c550 is unknown. The low midpoint redox potential (E m) of the purified soluble form (from −250 to −314mV) is incompatible with a redox function in PSII. However, more positive values for the Em have been obtained for the cyt c550 bound to the PSII. A very recent work has shown an E m value of +200mV. These data open the possibility of a redox function for this protein in electron transfer in PSII. Despite the long distance (22Å) between cyt c550 and the nearest redox cofactor (Mn4Ca cluster), an electron transfer reaction between these components is possible. Some kind of protective cycle involving a soluble redox component in the lumen has also been proposed. The aim of this article is to review previous studies done on cyt c550 and to consider its function in the light of the new results obtained in recent years. The emphasis is on the physical properties of the heme and its redox properties. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial. [Copyright &y& Elsevier]

Abstract: In dark-adapted plants and algae, chlorophyll a fluorescence induction peaks within 1s after irradiation due to well documented photochemical and non-photochemical processes. Here we show that the much slower fluorescence rise in cyanobacteria (the so-called “S to M rise” in tens of seconds) is due to state 2 to state 1 transition. This has been demonstrated in particular for Synechocystis PCC6803, using its RpaC− mutant (locked in state 1) and its wild-type cells kept in hyperosmotic suspension (locked in state 2). In both cases, the inhibition of state changes correlates with the disappearance of the S to M fluorescence rise, confirming its assignment to the state 2 to state 1 transition. The general physiological relevance of the SM rise is supported by its occurrence in several cyanobacterial strains: Synechococcus (PCC 7942, WH 5701) and diazotrophic single cell cyanobacterium (Cyanothece sp. ATCC 51142). We also show here that the SM fluorescence rise, and also the state transition changes are less prominent in filamentous diazotrophic cyanobacterium Nostoc sp. (PCC 7120) and absent in phycobilisome-less cyanobacterium Prochlorococcus marinus PCC 9511. Surprisingly, it is also absent in the phycobiliprotein rod containing Acaryochloris marina (MBIC 11017). All these results show that the S to M fluorescence rise reflects state 2 to state 1 transition in cyanobacteria with phycobilisomes formed by rods and core parts. We show that the pronounced SM fluorescence rise may reflect a protective mechanism for excess energy dissipation in those cyanobacteria (e.g. in Synechococcus PCC 7942) that are less efficient in other protective mechanisms, such as blue light induced non-photochemical quenching. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial. [Copyright &y& Elsevier]

Abstract: In order to prevent photodestruction by high light, photosynthetic organisms have evolved a number of mechanisms, known as non-photochemical quenching (NPQ), that deactivate the excited states of light harvesting pigments. Here we investigate the NPQ mechanism in the cyanobacterium Synechocystis sp. PCC 6803 mutant deficient in both photosystems. Using non-linear laser fluorimetry, we have determined molecular photophysical characteristics of phycocyanin and spectrally distinct forms of allophycocyanin for the cells in non-quenched and quenched states. Our analysis of non-linear fluorescence characteristics revealed that NPQ activation leads to an ~2-fold decrease in the relaxation times of both allophycocyanin fluorescence components, F660 and F680, and a 5-fold decrease in the effective excitation cross-section of F680, suggesting an emergence of a pathway of energy dissipation for both types of allophycocyanin. In contrast, NPQ does not affect the rates of singlet–singlet exciton annihilation. This indicates that, upon NPQ activation, the excess excitation energy is transferred from allophycocyanins to quencher molecules (presumably 3′hydroxyechinenone in the orange carotenoid protein), rather than being dissipated due to conformational changes of chromophores within the phycobilisome core. Kinetic measurements of fluorescence quenching in the Synechocystis mutant revealed the presence of several stages in NPQ development, as previously observed in the wild type. However, the lack of photosystems in the mutant enhanced the magnitude of NPQ as compared to the wild type, and allowed us to better characterize this process. Our results suggest a more complex kinetics of the NPQ process, thus clarifying a multistep model for the formation of the quenching center. [Copyright &y& Elsevier]

Abstract: Cyanobacteria respond to environmental stress conditions by adjusting their photosynthesis machinery. In Synechococcus sp. PCC 7942, phycobilisome degradation and other acclimation responses after nutrient or high light stress require activation by the phosphorylation-independent response regulator NblR. Structural modelling of its receiver domain suggested a role for Cys69 and Cys96 on activation of NblR. Here, we investigate this hypothesis by engineering Cys to Ala substitutions. In vivo and in vitro analyses indicated that mutations Cys69Ala and/or Cys96Ala have a minor impact on NblR function, structure, size, or oligomerization state of the protein, and that Cys69 and Cys96 do not seem to form disulphide bridges. Our results argue against the predicted involvement of Cys69 and Cys96 on NblR activation by redox sensing. [Copyright &y& Elsevier]

The percentage of spherical colonies from the trichomes of Nostoc sphaeroides reached 62-73% after 16 days with 50 and 250 μM P, but only10-15% at 0.5 and 5 μM P. During colony formation from microcolonies to macrocolonies, the growth rates were 95, 206 and 244% higher, respectively at 5, 50 and 250 μM P than that at 0.5 μM P. The light-saturated photosynthetic rate, maximum electron transport rate and light-limited photosynthetic efficiency at 0.5 μM P decreased, respectively, by 45, 51 and 32% than those at 250 μM P. These indicated that the colony development, growth and photosynthetic capacities were restricted at low P level, suggesting that P might be an important factor limiting the productivity and distribution of N. sphaeroides in the field. [ABSTRACT FROM AUTHOR]

Abstract: By using two strains of Arthrospira (Spirulina) platensis, an economically important filamentous cyanobacterium, we compared the impairment of PSII activity and loss of D1 protein content under UV-B radiation. Our study showed that UV-B radiation induced a gradual loss of the oxygen-evolving activity to about 56% after 180min UV-B irradiation both in strains 439 and D-0083, which have been kept under indoor and an outdoor culturing conditions, respectively for a prolonged period of time. The loss of oxygen evolution was accelerated in both strains in the presence of lincomycin, an inhibitor of protein synthesis, and the amount of D1 protein showed a decrease comparable to that of oxygen evolution during the UV-B exposure. However, the UV-B induced loss of oxygen-evolving activity and D1 protein amount was largely prevented when A. platensis cells were exposed to UV-B irradiance supplemented with visible light. Comparison of the two strains also showed a smaller extent of D1 protein synthesis dependent PSII repair in the indoor strain. Our results show that turnover of the D1 protein is an important defense mechanism to counteract the UV-B induced damage of PSII in A. platensis, and also that visible light plays an important role in maintaining the function of PSII under simultaneous exposure to UV-B and visible light. [Copyright &y& Elsevier]

Abstract: The production of high value biochemicals and high energy biofuels from sustainable resources through the use of microbial based, green conversion technologies could reduce the dependence on petrochemical resources. However, a sustainable source of carbon and a clean, cost effective method for its conversion to high quality biofuel products are obstacles that must be overcome. Here we describe the biosynthesis of fatty alcohols in a genetically engineered cyanobacterial system through heterologously expressing fatty acyl-CoA reductase and the effect of environmental stresses on the production of fatty alcohols in the mutant strains. Hydrocarbon production in three representative types of native cyanobacterial model strains and the mutant strain overexpressing acetyl-CoA carboxylase was evaluated. The results of this investigation demonstrate the potential for direct production of high value chemicals and high energy fuels in a single biological system that utilizes solar energy as the energy source and carbon dioxide as the carbon source. [Copyright &y& Elsevier]

We used differential scanning calorimetry (DSC) as a technique capable of identifying photosynthetic complexes on the basis of their calorimetric transitions. Annotation of thermal transitions was carried out with thylakoid membranes isolated from various photosynthetic mutants of Synechocystis sp. PCC6803. The thylakoid membranes exhibited seven major DSC bands between 40 and 85°C. The heat sorption curves were analyzed both by mathematical deconvolution of the overall endotherms and by a subsequent annealing procedure. The successive annealing procedure proved to be more reliable technique than mathematical deconvolution in assigning thermal transitions. The main DSC band, around 47°C, resulting from the high enthalpy change that corresponds to non-interacting complex of PSII, was assigned using the PSI-less/ apcE mutant cells. Another band around 68-70°C relates to the denaturation of PSII surrounded by other proteins of the photosynthetic complexes in wild type and PSI-less/ apcE cells. A further major transition found at 82-84°C corresponds to the PSI core complex of wild type and PSII-deficient BE cells. Other transition bands between 50-67 and 65-75°C are believed to relate to ATP synthase and cytochrome b f, respectively. These thermal transitions were obtained with thylakoids isolated from PSI/PSII mutant cells. Some minor bands determined at 59 and 83-84°C correspond to an unknown complex and NADH dehydrogenase, respectively. These annotations were done by PSI-less/ apcE and PSI/PSII mutants. [ABSTRACT FROM AUTHOR]

Summary: Anabaena sp. PCC7120 contains a gene, mrpA (all1838), which forms part of a seven gene-cluster (all1843–all1837) with significant sequence similarity to bacterial operons that putatively code for a multicomponent cation/proton antiporter involved in alkaline pH adaptation and salt resistance. We previously showed that growth and photosynthesis were inhibited in a strain mutated in mrpA, denoted as PHB11, particularly at alkaline pH. Here, we show that respiration was also impaired in the mutant independently of the external pH. In addition, at high pH, less ATP and vegetative cell ferredoxin were present in PHB11, which also showed lower levels of ferredoxin-NADP+ oxidoreductase (FNR). Ferredoxin and FNR are involved in the generation of reductant NADPH in cyanobacteria. These results suggest an energetic role of mrpA (and perhaps of the whole mrp-gene cluster) in Anabaena sp. PCC 7120 that is further supported by the significant similarity of putative Anabaena Mrp proteins to membrane subunits of complex I. [Copyright &y& Elsevier]

Abstract: Buoyancy provided by gas vesicles has been suggested to play an important role in regulating vertical distribution and nutrient acquisition in cyanobacteria. However, little is known about how changes in UV radiation (UVR, 280–400nm) would affect the buoyancy. We have shown here that the floatation activity of the economically important cyanobacterium Arthrospira platensis (D-0083) decreased with increased photosynthetic rates associated with increased photosynthetically active radiation (PAR), but it decreased less in the presence of UVR, which resulted in inhibitory effects. When the cells were grown under isoenergetic levels of solar PAR or UVR alone, they migrated downward under PAR but maintained buoyant under UVR. The buoyancy regulation of A. platensis depended on the exposed levels of PAR as well as UVR, which affected photosynthesis and growth in an antagonistic way. The buoyancy of A. platensis in water columns is likely to be dependant on diurnal photosynthetic performance regulated by solar radiation, and can hardly be considered as an active strategy to gain more energy during sunrise/sunset or to escape from harmful irradiation during the noon period. [Copyright &y& Elsevier]

Abstract: The physiological role of digalactosyldiacylglycerol (DGDG) in photosynthesis was examined using a dgdA mutant of Synechocystis sp. PCC 6803 that is defective in the biosynthesis of DGDG. The dgdA mutant cells showed normal growth under low light (LL) conditions. However, their growth was retarded under high light (HL) conditions and under Ca2+- and/or Cl−-limited conditions compared to wild-type cells. The retardation in growth of the mutant cells was recovered by exogenous supply of DGDG in the growth medium. The dgdA mutant showed increased sensitivity to photoinhibition. Although both photodamage and repair processes of photosynthesis were affected, the repair process was more severely affected than the photodamage process, suggesting that DGDG plays an important role in the photosynthetic repair cycle. [Copyright &y& Elsevier]

Abstract: Effects of two fertilizers, NH4Cl and KCl, on the growth of the edible cyanobacterium Ge–Xian–Mi (Nostoc) and four other cyanobacterial strains were compared at pH 8.3±0.2 and 25°C. Their growth was decreased by at least 65% at 10mmolL−1 NH4Cl but no inhibitory effect was observed at the same level of KCl. Meanwhile, the strains exhibited a great variation of sensitivity to NH4 + toxicity in the order: Ge–Xian–Mi> Anabaena azotica FACHB 118> Microcystis aeruginosa FACHB 905> M. aeruginosa FACHB 315> Synechococcus FACHB 805. The 96-h EC50 value for relative growth rate with regard to NH4 + for Ge–Xian–Mi was 1.105mmolL−1, which was much less than the NH4 + concentration in many agricultural soils (2–20mmolL−1). This indicated that the use of ammonium as nitrogen fertilizer was responsible for the reduced resource of Ge–Xian–Mi in the paddy field. After 96h exposure to 1mmolL−1 NH4Cl, the photosynthetic rate, F v/F m value, saturating irradiance for photosynthesis and PSII activity of Ge–Xian–Mi colonies were remarkably decreased. The chlorophyll synthesis of Ge–Xian–Mi was more sensitive to NH4 + toxicity than phycobiliproteins. Thus, the functional absorption cross section of Ge–Xian–Mi PSII was increased markedly at NH4Cl levels ≥1mmolL−1 and the electron transport on the acceptor side of PSII was significantly accelerated by NH4Cl addition ≥3mmolL−1. Dark respiration of Ge–Xian–Mi was significantly increased by 246% and 384% at 5 and 10mmolL−1 NH4Cl, respectively. The rapid fluorescence rise kinetics indicated that the oxygen-evolving complex of PSII was the inhibitory site of NH4 +. [Copyright &y& Elsevier]