LOPES, the LOFAR prototype station, was an antenna array for cosmic-ray air showers operating from 2003 to 2013 within the KASCADE-Grande experiment. Meanwhile, the analysis is finished and the data ...of air-shower events measured by LOPES are available with open access in the KASCADE Cosmic Ray Data Center (KCDC). This article intends to provide a summary of the achievements, results, and lessons learned from LOPES. By digital, interferometric beamforming the detection of air showers became possible in the radio-loud environment of the Karlsruhe Institute of Technology (KIT). As a prototype experiment, LOPES tested several antenna types, array configurations and calibration techniques, and pioneered analysis methods for the reconstruction of the most important shower parameters, i.e., the arrival direction, the energy, and mass-dependent observables such as the position of the shower maximum. In addition to a review and update of previously published results, we also present new results based on end-to-end simulations including all known instrumental properties. For this, we applied the detector response to radio signals simulated with the CoREAS extension of CORSIKA, and analyzed them in the same way as measured data. Thus, we were able to study the detector performance more accurately than before, including some previously inaccessible features such as the impact of noise on the interferometric cross-correlation beam. These results led to several improvements, which are documented in this paper and can provide useful input for the design of future cosmic-ray experiments based on the digital radio-detection technique.
Industrial fermentation in aerobic processes is plagued by high costs due to gas transfer limitations and substrate oxidation to CO
. It has been a longstanding challenge to engineer an obligate ...aerobe organism, such as
, into an anaerobe to facilitate its industrial application. However, the progress in this field is limited, due to the poor understanding of the constraints restricting its anoxic phenotype. In this paper, we provide a methodological description of a novel cultivation technology for
under anaerobic conditions, using the so-called microbial electrochemical technology within a bioelectrochemical system. By using an electrode as the terminal electron acceptor (mediated via redox chemicals), glucose catabolism could be activated without oxygen present. This (i) provides an anoxic-producing platform for sugar acid production at high yield and (ii) more importantly, enables systematic and quantitative characterizations of the phenotype of
in the absence of molecular oxygen. This unique electrode-based cultivation approach offers a tool to understand and in turn engineer the anoxic phenotype of
and possibly also other obligate aerobes.
Demosponges are a rich natural source of unusual lipids, some of which are of interest as geochemical biomarkers. Although demosponges are animals, they often host dense communities of microbial ...symbionts, and it is therefore unclear which lipids can be synthesized by the animal de novo, and which require input from the microbial community. To address this uncertainty, we analyzed the lipids of Amphimdeon queenslandica, the only demosponge with a published genome. We correlated the genetic and lipid repertoires of A. queenslandica to identify which biomarkers could potentially be synthesized and/or modified by the sponge. The fatty acid profile of A. queenslandica is dominated by an unusual Δ5,9 fatty acid (cis‐5,9‐hexacosadienoic acid)—similar to what has been found in other members of the Amphimdeon genus—while the sterol profile is dominated by C27‐C29 derivatives of cholesterol. Based on our analysis of the A. queenslandica genome, we predict that this sponge can synthesize sterols de novo, but it lacks critical genes necessary to synthesize basic saturated and unsaturated fatty acids. However, it does appear to have the genes necessary to modify simpler products into a more complex “algal‐like” assemblage of unsaturated fatty acids. Ultimately, our results provide additional support for the poriferan affinity of 24‐isopropylcholestanes in Neoproterozoic‐age rocks (the “sponge biomarker” hypothesis) and suggest that some algal proxies in the geochemical record could also have animal contributions.
There is currently a strong interest to derive the biological precursor cis,cis-muconic acid from shikimate pathway-branches to develop a biological replacement for adipic acid. Pioneered by the ...Frost laboratory this concept has regained interest: Recent approaches (Boles, Alper, Yan) however suffer from low product titres. Here an in silico comparison of all strain construction strategies was conducted to highlight stoichiometric optimizations. Using elementary mode analysis new knock-out strategies were determined in Saccharomyces cerevisiae and Escherichia coli. The strain construction strategies are unique to each pathway-branch and organism, allowing significantly different maximum and minimum yields. The maximum theoretical product carbon yields on glucose ranged from 86% (dehydroshikimate-branch) to 69% (anthranilate-branch). In most cases a coupling of product formation to growth was possible. Especially in S. cerevisiae chorismate-routes a minimum yield constraint of 46.9% could be reached. The knock-out targets are non-obvious, and not-transferable, highlighting the importance of tailored strain construction strategies.
•Maximum theoretical product carbon yields for biological production of cis,cis-muconic acid from glucose were calculated.•Existing strain construction strategies were compared in silico and weaknesses were identified.•New strain construction strategies were determined to allow coupling of product formation to growth for E. coli and S. cerevisiae.•In particular abolishing one target reaction introduces a minimum yield constraint of 28% in S. cerevisiae.•Determined knock-out targets are unique to each pathway and organism, highlighting the importance of tailored strain construction strategies.
Summary
In our modern ‘omics era, metabolic flux analysis (fluxomics) represents the physiological counterpart of its siblings transcriptomics, proteomics and metabolomics. Fluxomics integrates in ...vivo measurements of metabolic fluxes with stoichiometric network models to allow the determination of absolute flux through large networks of the central carbon metabolism. There are many approaches to implement fluxomics including flux balance analysis (FBA), 13C fluxomics and 13C‐constrained FBA as well as many experimental settings for flux measurement including dynamic, stationary and semi‐stationary. Here we outline the principles of the different approaches and their relative advantages. We demonstrate the unique contribution of flux analysis for phenotype elucidation using a thoroughly studied metabolic reaction as a case study, the microbial aerobic/anaerobic shift, highlighting the importance of flux analysis as a single layer of data as well as interlaced in multi‐omics studies.
A carbon‐free energy supply is essential to sustain our future. Biophotovoltaics (BPV) provides a promising solution for hydrogen supply by directly coupling light‐driven water splitting to hydrogen ...formation using oxygenic photoautotrophic cyanobacteria. However, BPV is currently limited by its low photon‐to‐current efficiency, and current experimental setups at a miniaturized scale hinder the rational investigation of the process and thus system optimization. In this article, we developed and optimized a new technical‐scale (~250 ml working volume) BPV platform with defined and controllable operating parameters. Factors that interfered with reproducible and stable current output signals were identified and adapted. We found that the classical BG11 medium, used for the cultivation of cyanobacteria and also in many BPV studies, caused severe interferences in the bioelectrochemical experiments. An optimized nBG11 medium guaranteed a low and stable background current in the BPV reactor, regardless of the presence of light and/or mediators. As proof‐of‐principle, a very high long‐term light‐dependent current output (peak current of over 20 µA) was demonstrated in the new set‐up over 12 days with living Synechocystis sp. PCC6803 cells and validated with appropriate controls. These results report the first reliable BPV platform generating reproducible photocurrent while still allowing quantitative investigation, rational optimization, and scale‐up of BPV processes.
A technical scale biophotovoltaics system designed for quantitative analysis of the cyanobacterial behaviors steered by the electrode was established and validated in this study. By implementing a systematically designed electrochemical‐compatible new BG11 (nBG11) medium, Lai and coworkers demonstrated a highly reproducible, long‐term stable and light‐associated electron flux from the model cyanobacteria Synechocystis sp. PCC6803 towards the anode via ferricyanide.
Microbial electrochemical technologies (MET) are promising to drive metabolic processes for the production of chemicals of interest. They provide microorganisms with an electrode as an electron sink ...or an electron source to stabilize their redox and/or energy state. Here, we applied an anode as additional electron sink to enhance the anoxic metabolism of the industrial bacterium Corynebacterium glutamicum through an anodic electro‐fermentation. In using ferricyanide as extracellular electron carrier, anaerobic growth was enabled and the feedback‐deregulated mutant Corynebacterium glutamicum lysC further accumulated L‐lysine. Under such oxidizing conditions we achieved L‐lysine titers of 2.9 mM at rates of 0.2 mmol/L/hr. That titer is comparable to recently reported L‐lysine concentrations achieved by anaerobic production under reductive conditions (cathodic electro‐fermentation). However unlike other studies, our oxidative conditions allowed anaerobic cell growth, indicating an improved cellular energy supply during anodic electro‐fermentation. In that light, we propose anodic electro‐fermentation as the right choice to support C. glutamicum stabilizing its redox and energy state and empower a stable anaerobic production of L‐lysine.
Growth and aerobic production of amino acids and various chemicals by important industrial bacterium, Corynebacterium glutamicum relies on oxygen as terminal electron acceptor, which limits product yields through substrate loss. To overcome the oxygen dependency, the authors provide the bacterium an alternative terminal electron acceptor, ferricyanide, which can be continuously re‐oxidized by an anode in a bioelectrochemical system. In that way, anodic electro‐fermentation enabled anaerobic growth, glucose consumption and production of organic acids and L‐lysine.
Mediator‐based extracellular electron transfer (EET) pathways can balance the redox metabolism of microbes. However, such electro‐biosynthesis processes are constrained by the unknown underlying EET ...mechanisms. In this paper, Pseudomonas putida was studied to systematically investigate its EET pathway to transition metal complexes (i. e., Fe(CN)63−/4− and Co(bpy)33+/2+; bpy=2,2′‐bipyridyl) under anaerobic conditions. Comparative proteomics showed the aerobic respiratory components were upregulated in a bioelectrochemical system without oxygen, suggesting their potential contribution to EET. Further tests found inhibiting cytochrome c oxidase activity by NaN3 and NADH dehydrogenase by rotenone did not significantly change the current output. However, the EET pathway was completely blocked, while cytochrome c reductase activity was inhibited by antimycin A. Although it cannot be excluded that cytochrome c and the periplasmic subunit of cytochrome c oxidase donate electrons to the transition metal complexes, these results strongly demonstrate that cytochrome c reductase is a key complex for the EET pathway.
Liquid wires for biohybrid systems: The electron transfer pathway between redox metal‐complexes and biological respiratory proteins is systematically investigated and revealed. Such a pathway can be readily applied to many non‐electrogenic microorganisms and thus expand the application of microbial electrochemical technology in many fields such as industrial biotechnology and biosensors.
Metabolite profiling technologies have improved to generate close to quantitative metabolomics data, which can be employed to quantitatively describe the metabolic phenotype of an organism. Here, we ...review the current technologies available for quantitative metabolomics, present their advantages and drawbacks, and the current challenges to generate fully quantitative metabolomics data. Metabolomics data can be integrated into metabolic networks using thermodynamic principles to constrain the directionality of reactions. Here we explain how to estimate Gibbs energy under physiological conditions, including examples of the estimations, and the different methods for thermodynamics-based network analysis. The fundamentals of the methods and how to perform the analyses are described. Finally, an example applying quantitative metabolomics to a yeast model by
C fluxomics and thermodynamics-based network analysis is presented. The example shows that (1) these two methods are complementary to each other; and (2) there is a need to take into account Gibbs energy errors. Better estimations of metabolic phenotypes will be obtained when further constraints are included in the analysis.
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