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2,002 results for: "biosynthesis" OR "adenosine triphosphate"
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  1. The effects of humic acid (HA) on interactions between ZnO nanoparticles (ZnO NPs) and Pseudomonas putida KT2440 biofilms at different maturity stages were investigated. Three stages of biofilm development were identified according to bacterial adenosine triphosphate (ATP) activity associated with biofilm development process. In the initial biofilm stage 1, the ATP content of bacteria was reduced by more than 90% when biofilms were exposed to ZnO NPs. But, in the mature biofilm stages 2 and 3, the ATP content was only slightly decreased. Biofilms at stage 3 exhibited less susceptibility to ZnO NPs than biofilms at stage 2. These resultsmore » suggest that more mature biofilms have a significantly higher tolerance to ZnO NPs compared to young biofilms. In addition, biofilms with intact extracellular polymeric substances (EPS) showed higher tolerance to ZnO NPs than those without EPS, indicating that EPS play a key role in alleviating the toxic effects of ZnO NPs. In both pure ZnO NPs and ZnO-HA mixtures, dissolved Zn 2+ originating from the NPs significantly contributed to the overall toxicity. The presence of HA dramatically decreased the toxicity of ZnO NPs due to the binding of Zn 2+ on HA. Furthermore, the combined results from this work suggest that the biofilm maturity stages and environmental constituents (such as humic acid) are important factors to consider when evaluating potential risks of NPs to ecological systems.« less
  2. Warming surface temperatures and increasing frequency and duration of widespread droughts threaten the health of natural forests and agricultural crops. High temperatures (HT) and intense droughts can lead to the excessive plant water loss and the accumulation of reactive oxygen species (ROS) resulting in extensive physical and oxidative damage to sensitive plant components including photosynthetic membranes. ROS signaling is tightly integrated with signaling mechanisms of the potent phytohormone abscisic acid (ABA), which stimulates stomatal closure leading to a reduction in transpiration and net photosynthesis, alters hydraulic conductivities, and activates defense gene expression including antioxidant systems. While generally assumed to bemore » produced in roots and transported to shoots following drought stress, recent evidence suggests that a large fraction of plant ABA is produced in leaves via the isoprenoid pathway. Through stomatal regulation and stress signaling which alters water and carbon fluxes, we highlight the fact that ABA lies at the heart of the Carbon-Water-ROS Nexus of plant response to HT and drought stress. We discuss the current state of knowledge of ABA biosynthesis, transport, and degradation and the role of ABA and other isoprenoids in the oxidative stress response. We discuss potential variations in ABA production and stomatal sensitivity among different plant functional types including isohydric/anisohydric and pioneer/climax tree species. We describe experiments that would demonstrate the possibility of a direct energetic and carbon link between leaf ABA biosynthesis and photosynthesis, and discuss the potential for a positive feedback between leaf warming and enhanced ABA production together with reduced stomatal conductance and transpiration. Finally, we propose a new modeling framework to capture these interactions. We conclude by discussing the importance of ABA in diverse tropical ecosystems through increases in the thermotolerance of photosynthesis to drought and heat stress, and the global importance of these mechanisms to carbon and water cycling under climate change scenarios.« less
  3. Methane utilization by methanotrophic bacteria is an attractive application for biotechnological conversion of natural or biogas into high-added-value products. Haloalcaliphilic methanotrophic bacteria belonging to the genus Methylomicrobium are among the most promising strains for methane-based biotechnology, providing easy and inexpensive cultivation, rapid growth, and the availability of established genetic tools. A number of methane bioconversions using these microbial cultures have been discussed, including the derivation of biodiesel, alkanes, and OMEGA-3 supplements. These compounds are derived from bacterial fatty acid pools. Here, we investigate fatty acid biosynthesis in Methylomicrobium buryatense 5G(B1). Most of the genes homologous to typical Type II fattymore » acid biosynthesis pathways could be annotated by bioinformatics analyses, with the exception of FA transport and regulatory elements. Different approaches for improving fatty acid accumulation were investigated. These studies indicated that both fatty acid degradation and acetyl- and malonyl-CoA levels are bottlenecks for higher level fatty acid production. The best strain generated in this study synthesizes 111 ± 2 mg/gDCW of extractable fatty acids, which is ~20% more than the original strain. A candidate gene for FA-biosynthesis regulation, farE, was identified and studied. Its deletion resulted in drastic changes to the FA profile, leading to an increased pool of C18-fatty acid methyl ester. The FarE-regulon was further investigated by RNA-seq analysis of gene expression in farE-knockout mutants and farE-overexpressing strains. These gene profiles highlighted a novel set of enzymes and regulators involved in fatty acid biosynthesis. As a result, the gene expression and fatty acid profiles of the different farE-strains support the hypothesis that metabolic fluxes upstream of fatty acid biosynthesis restrict fatty acid production in the methanotroph.« less
  4. With an aim to understand the origin and key contributing factors towards carboninduced cytotoxicity, we have studied five different carbon samples with diverse surface area, pore width, shape and size, conductivity and surface functionality. All the carbon materials were characterized with surface area and pore size distribution, x-ray photoelectron spectroscopy (XPS) and electron microscopic imaging. We performed cytotoxicity study in Caco-2 cells by colorimetric assay, oxidative stress analysis by reactive oxygen species (ROX) detection, cellular metabolic activity measurement by adenosine triphosphate (ATP) depletion and visualization of cellular internalization by TEM imaging. The carbon materials demonstrated a varying degree of cytotoxicitymore » in contact with Caco-2 cells. The lowest cell survival rate was observed for nanographene, which possessed the minimal size amongst all the carbon samples under study. None of the carbons induced oxidative stress to the cells as indicated by the ROX generation results. Cellular metabolic activity study revealed that the carbon materials caused ATP depletion in cells and nanographene caused the highest depletion. Visual observation by TEM imaging indicated the cellular internalization of nanographene. This study confirmed that the size is the key cause of carbon-induced cytotoxicity and it is probably caused by the ATP depletion within the cell.« less
  5. A multi-omics quantitative integrative analysis of lignin biosynthesis can advance the strategic engineering of wood for timber, pulp, and biofuels. Lignin is polymerized from three monomers (monolignols) produced by a grid-like pathway. The pathway in wood formation of Populus trichocarpa has at least 21 genes, encoding enzymes that mediate 37 reactions on 24 metabolites, leading to lignin and affecting wood properties. We perturb these 21 pathway genes and integrate transcriptomic, proteomic, fluxomic and phenomic data from 221 lines selected from ~2000 transgenics (6-month-old). The integrative analysis estimates how changing expression of pathway gene or gene combination affects protein abundance, metabolic-flux,more » metabolite concentrations, and 25 wood traits, including lignin, tree-growth, density, strength, and saccharification. The analysis then predicts improvements in any of these 25 traits individually or in combinations, through engineering expression of specific monolignol genes. The analysis may lead to greater understanding of other pathways for improved growth and adaptation.« less
  6. Here, the human pathogen Mycobacterium tuberculosis ( Mtb) requires a proteasome system to cause lethal infections in mice. We recently found that proteasome accessory factor E (PafE, Rv3780) activates proteolysis by the Mtb proteasome independently of adenosine triphosphate (ATP). Moreover, PafE contributes to the heat-shock response and virulence of Mtb. Here, we show that PafE subunits formed four-helix bundles similar to those of the eukaryotic ATP-independent proteasome activator subunits of PA26 and PA28. However, unlike any other known proteasome activator, PafE formed dodecamers with 12-fold symmetry, which required a glycine-XXX-glycine-XXX-glycine motif that is not found in previously described activators. Intriguingly,more » the truncation of the PafE carboxyl-terminus resulted in the robust binding of PafE rings to native proteasome core particles and substantially increased proteasomal activity, suggesting that the extended carboxyl-terminus of this cofactor confers suboptimal binding to the proteasome core particle. Collectively, our data show that proteasomal activation is not limited to hexameric ATPases in bacteria.« less
  7. Mycobacterium tuberculosis encodes a proteasome that is highly similar to eukaryotic proteasomes and is required to cause lethal infections in animals. The only pathway known to target proteins for proteasomal degradation in bacteria is pupylation, which is functionally analogous to eukaryotic ubiquitylation. However, evidence suggests that the M. tuberculosis proteasome contributes to pupylation-independent pathways as well. To identify new proteasome cofactors that might contribute to such pathways, we isolated proteins that bound to proteasomes overproduced in M. tuberculosis and found a previously uncharacterized protein, Rv3780, which formed rings and capped M. tuberculosis proteasome core particles. Rv3780 enhanced peptide and proteinmore » degradation by proteasomes in an adenosine triphosphate (ATP)-independent manner. We identified putative Rv3780-dependent proteasome substrates and found that Rv3780 promoted robust degradation of the heat shock protein repressor, HspR. Importantly, an M. tuberculosis Rv3780 mutant had a general growth defect, was sensitive to heat stress, and was attenuated for growth in mice. Collectively, these data demonstrate that ATP-independent proteasome activators are not confined to eukaryotes and can contribute to the virulence of one the world’s most devastating pathogens.« less
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  8. Here, living systems possess a rich biochemistry that can be harnessed through metabolic engineering to produce valuable therapeutics, fuels and fine chemicals. In spite of the tools created for this purpose, many organisms tend to be recalcitrant to modification or difficult to optimize. Crude cellular extracts, made by lysis of cells, possess much of the same biochemical capability, but in an easier to manipulate context. Metabolic engineering in crude extracts, or cell-free metabolic engineering, can harness these capabilities to feed heterologous pathways for metabolite production and serve as a platform for pathway optimization. However, the inherent biochemical potential of amore » crude extract remains ill-defined, and consequently, the use of such extracts can result in inefficient processes and unintended side products. Herein, we show that changes in cell growth conditions lead to changes in the enzymatic activity of crude cell extracts and result in different abilities to produce the central biochemical precursor pyruvate when fed glucose. Proteomic analyses coupled with metabolite measurements uncover the diverse biochemical capabilities of these different crude extract preparations and provide a framework for how analytical measurements can be used to inform and improve crude extract performance. Such informed developments can allow enrichment of crude extracts with pathways that promote or deplete particular metabolic processes and aid in the metabolic engineering of defined products.« less

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