Photoproduction of Hydrogen by the Marine Heterocystous Cyanobacterium Anabaena Species TU37-1 Under a Nitrogen Atmosphere

Hydrogen production rates by Anabaena sp. strain TU37-1 obtained after an initial 1-day incubation period were approximately 70 to 80 and 3 to 9 µmol (mg chl)^–1 h^–1 under argon and nitrogen atmospheres, respectively. Hydrogen production under argon was not enhanced by addition of carbon dioxide, but was enhanced to some extent under nitrogen by increasing the initial carbon dioxide concentration. Rates of hydrogen and oxygen production during the initial 7-hour period were 15 and 220 µmol (mg chl)^–1 h^–1, respectively, in vessels with 18.5% initial carbon dioxide. Hydrogen production under nitrogen was enhanced by addition of carbon monoxide (1%). The rate obtained from the initial 1-day incubation period was about 40 µmol (mg chl)^–1 h^–1, which corresponded to about 60% of that under argon. On the basis of these observations, a possible strategy for hydrogen production by nitrogen-fixing cyanobacteria under nitrogen in the presence of carbon monoxide is indicated.     

Pathways of Carbon Assimilation and Ammonia Oxidation Suggested by Environmental Genomic Analyses of Marine Crenarchaeota

Marine Crenarchaeota represent an abundant component of oceanic microbiota with potential to significantly influence biogeochemical cycling in marine ecosystems. Prior studies using specific archaeal lipid biomarkers and isotopic analyses indicated that planktonic Crenarchaeota have the capacity for autotrophic growth, and more recent cultivation studies support an ammonia-based chemolithoautotrophic energy metabolism. We report here analysis of fosmid sequences derived from the uncultivated marine crenarchaeote, Cenarchaeum symbiosum, focused on the reconstruction of carbon and energy metabolism. Genes predicted to encode multiple components of a modified 3-hydroxypropionate cycle of autotrophic carbon assimilation were identified, consistent with utilization of carbon dioxide as a carbon source. Additionally, genes predicted to encode a near complete oxidative tricarboxylic acid cycle were also identified, consistent with the consumption of organic carbon and in the production of intermediates for amino acid and cofactor biosynthesis. Therefore, C. symbiosum has the potential to function either as a strict autotroph, or as a mixotroph utilizing both carbon dioxide and organic material as carbon sources. From the standpoint of energy metabolism, genes predicted to encode ammonia monooxygenase subunits, ammonia permease, urease, and urea transporters were identified, consistent with the use of reduced nitrogen compounds as energy sources fueling autotrophic metabolism. Homologues of these genes, recovered from ocean waters worldwide, demonstrate the conservation and ubiquity of crenarchaeal pathways for carbon assimilation and ammonia oxidation. These findings further substantiate the likely global metabolic importance of Crenarchaeota with respect to key steps in the biogeochemical transformation of carbon and nitrogen in marine ecosystems.     

Inhibition of Nitrogen and Photosynthetic Carbon Assimilation of Maize Seedlings by Exposure to a Combination of Salt Stress and Potassium-Deficient Stress

The main aim of this work is to identify how the combined stresses affect the interdependent nitrogen and photosynthetic carbon assimilations in maize. Maize plants were cultivated in Meider's solution. They were subjected to salt stress and potassium deficiency in the K-present Meider's media and K-deficient Meider's media. After 5?weeks, we measured chlorophyll a fluorescence and the activities of several enzymes in metabolic checkpoints coordinating primary nitrogen and carbon assimilation in the leaves of maize. The study showed that the combination of salt stress and potassium-deficient stress more significantly decreased nitrate uptake, plant growth, the activities of nitrate reductase, glutamate dehydrogenase, glutamate synthase, urease, glutamic-pyruvic transaminase, glutamic-oxaloace transaminase, sucrose-phosphate synthase, phosphoenolpyruvate carboxylase, and the synthesis of free amino acids, chlorophyll, and protein than those of each individual stress, respectively. However, the combined stresses significantly increased the accumulation of ammonium and carbohydrate products. The combined stresses also significantly decreased the oxygen evolution, the electron transport, and the efficiency of photochemical energy conversion by photosystem II in maize seedlings. Taken together, a combination of salt stress and potassium-deficient stress impaired the assimilations of both nitrogen and carbon and decreased the photosystem II activity in maize.     

Long-term Changes in Forest Carbon and Nitrogen Cycling Caused by an Introduced Pest/Pathogen Complex

Invasion of exotic forest pests and pathogens is a serious environmental problem for many forests throughout the world, and has been especially damaging to forests of eastern North America. We studied the impacts of an exotic pest/pathogen complex, the beech bark disease (BBD), in the Catskill Mountains of New York State, USA. In this region, BBD has caused a decline in the basal area of American beech (Fagus grandifolia Ehrh.) over the last 60 years and this decline has been accompanied by an increase in the basal area of sugar maple (Acer saccharum Marsh.). We studied the impacts of the BBD on carbon (C) and nitrogen (N) cycling using a series of stands that represented a sequence of disease impact and beech replacement by sugar maple. Our study showed that these long-term changes in tree species composition can lead to important changes in C and N cycling in the ecosystem, including an increase in litter decomposition, a decrease in soil C:N ratio, and an increase in extractable nitrate in the soil and nitrate in soil solution. Rates of potential net N mineralization and nitrification did not change across the BBD sequence, but the fraction of mineralized N that was nitrified increased significantly. Many of the observed changes in ecosystem function are larger in magnitude than those attributed to climate change or air pollution, suggesting that the impacts of invasive pests and pathogens on tree species composition could be one of the most important factors driving changes in C and N cycling in these forests in the coming decades.     

Spatial Perturbations within an RNA Promoter Specifically Recognized by a Viral RNA-Dependent RNA Polymerase (RdRp) Reveal That RdRp Can Adjust Its Promoter Binding Sites

RNA synthesis during viral replication requires specific recognition of RNA promoters by the viral RNA-dependent RNA polymerase (RdRp). Four nucleotides (−17, −14, −13, and −11) within the brome mosaic virus (BMV) subgenomic core promoter are required for RNA synthesis by the BMV RdRp (R. W. Siegel et al., Proc. Natl. Acad. Sci. USA 94:11238–11243, 1997). The spatial requirements for these four nucleotides and the initiation (+1) cytidylate were examined in RNAs containing nucleotide insertions and deletions within the BMV subgenomic core promoter. Spatial perturbations between nucleotides −17 and −11 resulted in decreased RNA synthesis in vitro. However, synthesis was still dependent on the key nucleotides identified in the wild-type core promoter and the initiation cytidylate. In contrast, changes between nucleotides −11 and +1 had a less severe effect on RNA synthesis but resulted in RNA products initiated at alternative locations in addition to the +1 cytidylate. The results suggest a degree of flexibility in the recognition of the subgenomic promoter by the BMV RdRp and are compared with functional regions in other DNA and RNA promoters.     

Evidence for an Ionic Intermediate in the Transformation of Fatty Acid Hydroperoxide by a Catalase-related Allene Oxide Synthase from the Cyanobacterium Acaryochloris marina

Allene oxides are reactive epoxides biosynthesized from fatty acid hydroperoxides by specialized cytochrome P450s or by catalase-related hemoproteins. Here we cloned, expressed, and characterized a gene encoding a lipoxygenase-catalase/peroxidase fusion protein from Acaryochloris marina. We identified novel allene oxide synthase (AOS) activity and a by-product that provides evidence of the reaction mechanism. The fatty acids 18.4ω3 and 18.3ω3 are oxygenated to the 12R-hydroperoxide by the lipoxygenase domain and converted to the corresponding 12R,13-epoxy allene oxide by the catalase-related domain. Linoleic acid is oxygenated to its 9R-hydroperoxide and then, surprisingly, converted ∼70% to an epoxyalcohol identified spectroscopically and by chemical synthesis as 9R,10S-epoxy-13S-hydroxyoctadeca-11E-enoic acid and only ∼30% to the 9R,10-epoxy allene oxide. Experiments using oxygen-18-labeled 9R-hydroperoxide substrate and enzyme incubations conducted in H₂¹⁸O indicated that ∼72% of the oxygen in the epoxyalcohol 13S-hydroxyl arises from water, a finding that points to an ionic intermediate (epoxy allylic carbocation) during catalysis. AOS and epoxyalcohol synthase activities are mechanistically related, with a reacting intermediate undergoing a net hydrogen abstraction or hydroxylation, respectively. The existence of epoxy allylic carbocations in fatty acid transformations is widely implicated although for AOS reactions, without direct experimental support. Our findings place together in strong association the reactions of allene oxide synthesis and an ionic reaction intermediate in the AOS-catalyzed transformation.A diverse spectrum of signaling molecules is biosynthesized in nature from polyunsaturated fatty acids, their peroxides, and further transformations of the fatty acid peroxides. The peroxides are formed by two classes of dioxygenase enzyme. The hemoprotein dioxygenases include prostaglandin H synthase (cyclooxygenase) in animals (1), α-dioxygenase in plants (2), and several linoleate dioxygenases in fungi (3–5). The non-heme iron lipoxygenases are even more widespread, being almost ubiquitous among organisms that contain polyunsaturated fatty acids (6–8). Although further biosynthetic transformation is sometimes accomplished by an additional catalytic activity of the initiating dioxygenase (e.g. leukotriene A₄ synthase (9) or aldehyde-synthesizing hydroperoxide cleaving activity (10) among the LOX² enzymes), commonly another distinct enzyme is used to rearrange or otherwise modify the reactive fatty acid peroxide intermediate. Two hemoprotein types are found that have become specialized for this biosynthetic role: cytochrome P450s and catalase-related enzymes.The fatty acid peroxide-metabolizing P450s are by far the better known and include CYP5 (thromboxane synthase) and CYP8A (prostacyclin synthase) in animals (11), and the entire family of CYP74 in plants (12). The individual CYP74 enzymes include allene oxide synthase (AOS), one of which catalyzes a key step in cyclopentenone synthesis in the jasmonate pathway, hydroperoxide lyase, divinyl ether synthase, and epoxyalcohol synthase (12, 13). The catalase-related enzymes are distinctive in that they have been found naturally as a fusion protein with the LOX enzyme that forms their hydroperoxide substrate (14). The known activities include AOS in Plexaura homomalla and other marine corals (with a different specificity for fatty acid hydroperoxide compared with the plant P450 AOS) (15, 16), and the unique bicyclobutane synthase and other allylic epoxide synthase activities of the enzyme in the cyanobacterium Anabaena PCC-7120 (17, 18). Currently we are trying to understand the scope of the reactions catalyzed by this catalase-related family of enzymes. Our objectives are to help understand the structure-function relationships in the reactions with peroxides, to provide new insights on the mechanism of these hemoprotein-catalyzed transformations, as well as, by reflection, to give a different perspective on the parent hemoprotein, the hydrogen peroxide-metabolizing true catalase.The underlying chemistry of the fatty acid hydroperoxide transformations by the specialized P450 and catalase-related hemoproteins can be written as purely free radical in character, or ionic, or with facets of both (19–22). Reaction is generally considered to be initiated by homolytic cleavage of the peroxide O–O bond (see Fig. 1). While subsequent reactions can be construed as following a radical pathway to product (e.g. in the hydroperoxide lyase reaction (19)), others are considered to involve an electron transfer step, giving a carbocation intermediate in the penultimate steps to product (22) (Fig. 1). The underlying grounds for these mechanisms are more a subject of debate and of comparison to other chemistry than of defining evidence on the reactions in question. The results reported here add a measure of experimental support for ionic events in the peroxide transformation by a catalase-related hemoprotein.Open in a separate windowFIGURE 1.Hemoprotein-catalyzed transformation of fatty acid hydroperoxides via diverging routes. A radical pathway can lead to aldehydes (left side) and an ionic pathway via a putative carbocation intermediate to allene oxides (right side).Recently genome sequencing was completed on the cyanobacterium Acaryochloris marina (23). A. marina is a focus of attention owing to its harboring a light-harvesting complex containing the unusual chlorophyll d, considered a bridge in the evolutionary development of photosynthetic mechanisms (24, 25). BLAST searches of the A. marina genome reveal three individual LOX sequences and additionally, the presence of DNA encoding a putative fusion protein of lipoxygenase and catalase-related hemoprotein, the topic of this report. We find an unexpected by-product in the reaction with one particular fatty acid (C18.2ω6); the partial incorporation of ¹⁸O from water in a newly formed hydroxyl group has implications related to the mechanism of hydroperoxide transformation.     

A Polyaniline‐Coated Sulfur/Carbon Composite with an Enhanced High‐Rate Capability as a Cathode Material for Lithium/Sulfur Batteries

Polyaniline‐coated sulfur/conductive‐carbon‐black (PANI@S/C) composites with different contents of sulfur are prepared via two facile processes including ball‐milling and thermal treatment of the conductive carbon black and sublimed sulfur, followed by an in situ chemical oxidative polymerization of the aniline monomer in the presence of the S/C composite and ammonium persulfate. The microstructure and electrochemical performance of the as‐prepared composites are investigated systematically. It is demonstrated that the polyaniline, with a thickness of ≈5–10 nm, is coated uniformly onto the surface of the S/C composite forming a core/shell structure. The PANI@S/C composite with 43.7 wt% sulfur presents the optimum electrochemical performance, including a large reversible capacity, a good coulombic efficiency, and a high active‐sulfur utilization. The formation of the unique core/shell structure in the PANI@S/C composites is responsible for the improvement of the electrochemical performance. In particular, the high‐rate charge/discharge capability of the PANI@S/C composites is excellent due to a synergistic effect on the high electrical conductivity from both the conductive carbon black in the matrix and the PANI on the surface. Even at an ultrahigh rate (10C), a maximum discharge capacity of 635.5 mA h per g of sulfur is still retained for the PANI@S/C composite after activation, and the discharge capacity retention is over 60% after 200 cycles.     

Photoassimilation of CO2 by Isolated Bundle-Sheath Strands of Zea mays I. Stimulation of CO2 Assimilation by Adding Various Intermediates of the Photosynthetic Cycle; Evidence for a Deficient Photosystem II Activity

Isolated bundle-sheath (BS) strands from leaves of mature maize plants show enhanced rates of CO₂ fixation in the presence of reduced intermediates of the photosynthetic cycle (R5P, DHAP, FruDP.) 3PGA is the major labelled product of ¹⁴CO₂ fixation whatever the substrate added. CO₂ fixation is much lower with PGA than with reduced intermediates, suggesting a limited capacity of the cells to regenerate RuDP (the CO₂-acceptor) from PGA. These two experimental facts, which are characteristic features of bundle-sheath photosynthesis for maize (a species with agranal bundle-sheath chloroplasts) indicate that phaotosystem II activity is a limiting factor for the evolution of the bundle-sheath photosynthetic process. Nevertheless, a reducing capacity arises as proved by sensitivity of CO₂ fixation to DCMU, particularly when PGA is added to the bundle-sheath. PGA synthesis occurs, in the presence of non-limiting amounts of CO₂, according to the equation: RuDP + CO₂→ 2 PGA; the oxygen effect on ¹⁴CO₂ fixation, at lower CO₂ concentration, is interpreted as a dilution effect of the internal pool of ¹⁴CO₂ by unlabelled CO₂ generated by photorespiration.     

Strategies for Primary Prevention of Coronary Heart Disease Based on Risk Stratification by the ACC/AHA Lipid Guidelines,ATP III Guidelines,Coronary Calcium Scoring,and C-Reactive Protein,and a Global Treat-All Strategy: A Comparative--Effectiveness Modeling Study

Background

Several approaches have been proposed for risk-stratification and primary prevention of coronary heart disease (CHD), but their comparative and cost-effectiveness is unknown.

Methods

We constructed a state-transition microsimulation model to compare multiple approaches to the primary prevention of CHD in a simulated cohort of men aged 45–75 and women 55–75. Risk-stratification strategies included the 2013 American College of Cardiology/American Heart Association (ACC/AHA) guidelines on the treatment of blood cholesterol, the Adult Treatment Panel (ATP) III guidelines, and approaches based on coronary artery calcium (CAC) scoring and C-reactive protein (CRP). Additionally we assessed a treat-all strategy in which all individuals were prescribed either moderate-dose or high-dose statins and all males received low-dose aspirin. Outcome measures included CHD events, costs, medication-related side effects, radiation-attributable cancers, and quality-adjusted-life-years (QALYs) over a 30-year timeframe.

Results

Treat-all with high-dose statins dominated all other strategies for both men and women, gaining 15.7 million QALYs, preventing 7.3 million myocardial infarctions, and saving over $238 billion, compared to the status quo, far outweighing its associated adverse events including bleeding, hepatitis, myopathy, and new-onset diabetes. ACC/AHA guidelines were more cost-effective than ATP III guidelines for both men and women despite placing 8.7 million more people on statins. For women at low CHD risk, treat-all with high-dose statins was more likely to cause a statin-related adverse event than to prevent a CHD event.

Conclusions

Despite leading to a greater proportion of the population placed on statin therapy, the ACC/AHA guidelines are more cost-effective than ATP III. Even so, at generic prices, treating all men and women with statins and all men with low-dose aspirin appears to be more cost-effective than all risk-stratification approaches for the primary prevention of CHD. Especially for low-CHD risk women, decisions on the appropriate primary prevention strategy should be based on shared decision making between patients and healthcare providers.     

Location of the structural gene for glycyl ribonucleic acid synthetase by means of a strain ofEscherichia coli possessing an unusual enzyme

Summary E. coli KB (Benzer) differs from other common laboratory strains in possessing a glycyl sRNA synthetase with a 50 to 100 times elevated K _m for glycine. The degree of charging of glycyl sRNA in this strain can be increased by supplementing the growth medium with glycine. The altered enzyme has been used as a marker by which to map its structural gene. Linkage analysis of recombinants from uninterrupted matings, and cotransduction (80%) of the synthetase withxyl, indicate that this gene is located betweenxyl andmalt, close toxyl, at min 69.5 on the map drawn byTaylor andThoman (1964).     

One-step process for debromination and aerobic mineralization of tetrabromobisphenol-A by a novel Ochrobactrum sp. T isolated from an e-waste recycling site

A novel bacterium, Ochrobactrum sp. T, capable of simultaneous debromination and aerobic mineralization of tetrabromobisphenol-A (TBBPA), was isolated from a sludge sample collected from an electronic-waste recycling site. The bacterium exhibited maximal debrominase activity at pH 6.5, 35 °C, and 200 rpm in Luria-Bertani culture medium. Initial TBBPA concentration and pH had more significant effects on degradation efficiency than those of temperature and inoculum size. Degradation and debromination efficiencies of 91.8% and 86.7%, respectively, were achieved within 72 h under optimized conditions of 35 °C, pH 7.0, inoculum volume of 25 mL, and TBBPA concentration of 3 mg L⁻¹. In addition, a 35.6% decrease in total organic carbon was observed after the degradation of 5 mg L⁻¹ TBBPA for 120 h. Eight metabolic intermediates were identified during the biodegradation of TBBPA. This study is the first report to propose a one-step process for TBBPA debromination and mineralization by a single bacterial strain.     

Validation study of a method for assaying DE-310, a macromolecular carrier conjugate containing an anti-tumor camptothecin derivative, and the free drug in human plasma by HPLC and LC/MS/MS

DE-310 is a macromolecular carrier conjugate containing an anti-tumor camptothecin derivative, DX-8951, conjugated to a water-soluble polymer by means of a peptide spacer. New assay methods have been developed to determine the polymer-bonded DX-8951 conjugate, free DX-8951, and Glycyl-DX-8951 in human plasma. Solid-phase extraction was used to extract free DX-8951 and Glycyl-DX-8951 from plasma, and LC/MS/MS (Method I) was used to determine the amount of each analyte. Protein precipitation was used to extract Conjugated DX-8951, which was then digested with thermolysin. HPLC (Method II) was used to determine the productive compound (Phenylalanyl-Glycyl-DX-8951). The lower limit of quantitation of DX-8951 was 50 pg/ml, of Glycyl-DX-8951 was 80 pg/ml, and of Conjugated DX-8951 was 100 ng/ml (as DX-8951 equivalent). Both methods showed satisfactory sensitivity, precision, and accuracy.     

Potential Global Standard Stratotype-section and Point (GSSP) for a Cambrian stage boundary defined by the first appearance of the trilobite Ptychagnostus atavus, Drum Mountains, Utah, USA

The base of the Ptychagnostus (or Acidusus) atavus Zone is one of the most clearly recognizable horizons on an intercontinental scale in the Cambrian System, and would serve as an excellent position for the base of a new stage-level chronostratigraphic subdivision. Among well-exposed, readily accessible sections in Laurentia, the “Stratotype Ridge” section, Drum Mountains, western Utah, USA, is perhaps the most suitable for a Global Standard Stratotype-section and Point (GSSP) defined by the first appearance datum (FAD) of the cosmopolitan agnostoid trilobite P. atavus. In the “Stratotype Ridge” section, the FAD of P. atavus occurs near the base of a calcisiltite bed 62 m above the base of the Wheeler Formation. A position corresponding closely to this horizon can be recognized with precision in Gondwana, Siberia, Kazakhstan, and Baltica using a combination of stratigraphic tools, the most useful of which are trilobite biostratigraphy, conodont biostratigraphy, and sequence stratigraphy. Brachiopod biostratigraphy and chemostratigraphy provide general constraints on the position of the horizon intercontinentally.