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Start date: 2015 × Subject: metabolism ×

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Now showing 1 - 10 of 26
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    Detection of Protein Glycosylation Using Tip-Enhanced Raman Scattering
    (Columbus, Ohio : American Chemical Society, 2016) Cowcher, David P.; Deckert-Gaudig, Tanja; Brewster, Victoria L.; Ashton, Lorna; Deckert, Volker; Goodacre, Royston
    The correct glycosylation of biopharmaceutical glycoproteins and their formulations is essential for them to have the desired therapeutic effect on the patient. It has recently been shown that Raman spectroscopy can be used to quantify the proportion of glycosylated protein from mixtures of native and glycosylated forms of bovine pancreatic ribonuclease (RNase). Here we show the first steps toward not only the detection of glycosylation status but the characterization of glycans themselves from just a few protein molecules at a time using tip-enhanced Raman scattering (TERS). While this technique generates complex data that are very dependent on the protein orientation, with the careful development of combined data preprocessing, univariate and multivariate analysis techniques, we have shown that we can distinguish between the native and glycosylated forms of RNase. Many glycoproteins contain populations of subtly different glycoforms; therefore, with stricter orientation control, we believe this has the potential to lead to further glycan characterization using TERS, which would have use in biopharmaceutical synthesis and formulation research.
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    PH-Responsive Biohybrid Carrier Material for Phenol Decontamination in Wastewater
    (Columbus, Ohio : American Chemical Soc., 2018) Pretscher, Martin; Pineda-Contreras, Beatriz A.; Kaiser, Patrick; Reich, Steffen; Schöbel, Judith; Kuttner, Christian; Freitag, Ruth; Fery, Andreas; Schmalz, Holger; Agarwal, Seema
    Smart polymers are a valuable platform to protect and control the activity of biological agents over a wide range of conditions, such as low pH, by proper encapsulation. Such conditions are present in olive oil mill wastewater with phenol as one of the most problematic constituents. We show that elastic and pH-responsive diblock copolymer fibers are a suitable carrier for Corynebacterium glutamicum, i.e., bacteria which are known for their ability to degrade phenol. Free C. glutamicum does not survive low pH conditions and fails to degrade phenol at low pH conditions. Our tea-bag like biohybrid system, where the pH-responsive diblock copolymer acts as a protecting outer shell for the embedded bacteria, allows phenol degradation even at low pH. Utilizing a two-step encapsulation process, planktonic cells were first encapsulated in poly(vinyl alcohol) to protect the bacteria against the organic solvents used in the second step employing coaxial electrospinning.
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    High-throughput screening Raman microspectroscopy for assessment of drug-induced changes in diatom cells
    (Cambridge : Royal Society of Chemistry, 2019) Rüger J.; Mondol A.S.; Schie I.W.; Popp J.; Krafft C.
    High-throughput screening Raman spectroscopy (HTS-RS) with automated localization algorithms offers unsurpassed speed and sensitivity to investigate the effect of dithiothreitol on the diatom Phaedactylum tricornutum. The HTS-RS capability that was demonstrated for this model system can be transferred to unmet analytical applications such as kinetic in vivo studies of microalgal assemblages. © 2019 The Royal Society of Chemistry.
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    Key knowledge and data gaps in modelling the influence of CO2 concentration on the terrestrial carbon sink
    (München : Elsevier, 2016) Pugh, T.A.M.; Müller, C.; Arneth, A.; Haverd, V.; Smith, B.
    Primary productivity of terrestrial vegetation is expected to increase under the influence of increasing atmospheric carbon dioxide concentrations ([CO2]). Depending on the fate of such additionally fixed carbon, this could lead to an increase in terrestrial carbon storage, and thus a net terrestrial sink of atmospheric carbon. Such a mechanism is generally believed to be the primary global driver behind the observed large net uptake of anthropogenic CO2 emissions by the biosphere. Mechanisms driving CO2 uptake in the Terrestrial Biosphere Models (TBMs) used to attribute and project terrestrial carbon sinks, including that from increased [CO2], remain in large parts unchanged since those models were conceived two decades ago. However, there exists a large body of new data and understanding providing an opportunity to update these models, and directing towards important topics for further research. In this review we highlight recent developments in understanding of the effects of elevated [CO2] on photosynthesis, and in particular on the fate of additionally fixed carbon within the plant with its implications for carbon turnover rates, on the regulation of photosynthesis in response to environmental limitations on in-plant carbon sinks, and on emergent ecosystem responses. We recommend possible avenues for model improvement and identify requirements for better data on core processes relevant to the understanding and modelling of the effect of increasing [CO2] on the global terrestrial carbon sink.
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    Production of highly concentrated and hyperpolarized metabolites within seconds in high and low magnetic fields
    (Cambridge : RSC Publ., 2019) Korchak, Sergey; Emondts, Meike; Mamone, Salvatore; Blümich, Bernhard; Glöggler, Stefan
    Hyperpolarized metabolites are very attractive contrast agents for in vivo magnetic resonance imaging studies enabling early diagnosis of cancer, for example. Real-time production of concentrated solutions of metabolites is a desired goal that will enable new applications such as the continuous investigation of metabolic changes. To this end, we are introducing two NMR experiments that allow us to deliver high levels of polarization at high concentrations (50 mM) of an acetate precursor (55% 13C polarization) and acetate (17% 13C polarization) utilizing 83% para-state enriched hydrogen within seconds at high magnetic field (7 T). Furthermore, we have translated these experiments to a portable low-field spectrometer with a permanent magnet operating at 1 T. The presented developments pave the way for a rapid and affordable production of hyperpolarized metabolites that can be implemented in e.g. metabolomics labs and for medical diagnosis.
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    Monoclonal Antibodies 13A4 and AC133 Do Not Recognize the Canine Ortholog of Mouse and Human Stem Cell Antigen Prominin-1 (CD133)
    (San Francisco, California, US : PLOS, 2016) Thamm, Kristina; Graupner, Sylvi; Werner, Carsten; Huttner, Wieland B.; Corbeil, Denis; Nabi, Ivan R
    The pentaspan membrane glycoprotein prominin-1 (CD133) is widely used in medicine as a cell surface marker of stem and cancer stem cells. It has opened new avenues in stem cell-based regenerative therapy and oncology. This molecule is largely used with human samples or the mouse model, and consequently most biological tools including antibodies are directed against human and murine prominin-1. Although the general structure of prominin-1 including its membrane topology is conserved throughout the animal kingdom, its primary sequence is poorly conserved. Thus, it is unclear if anti-human and -mouse prominin-1 antibodies cross-react with their orthologs in other species, especially dog. Answering this issue is imperative in light of the growing number of studies using canine prominin-1 as an antigenic marker. Here, we address this issue by cloning the canine prominin-1 and use its overexpression as a green fluorescent protein fusion protein in Madin-Darby canine kidney cells to determine its immunoreactivity with antibodies against human or mouse prominin-1. We used immunocytochemistry, flow cytometry and immunoblotting techniques and surprisingly found no cross-species immunoreactivity. These results raise some caution in data interpretation when anti-prominin-1 antibodies are used in interspecies studies.
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    Redox Stimulation of Human THP-1 Monocytes in Response to Cold Physical Plasma
    (Austin, Tex. : Landes Bioscience, 2015) Bekeschus, Sander; Schmidt, Anke; Bethge, Lydia; Masur, Kai; von Woedtke, Thomas; Hasse, Sybille; Wende, Kristian
    In plasma medicine, cold physical plasma delivers a delicate mixture of reactive components to cells and tissues. Recent studies suggested a beneficial role of cold plasma in wound healing. Yet, the biological processes related to the redox modulation via plasma are not fully understood. We here used the monocytic cell line THP-1 as a model to test their response to cold plasma in vitro. Intriguingly, short term plasma treatment stimulated cell growth. Longer exposure only modestly compromised cell viability but apparently supported the growth of cells that were enlarged in size and that showed enhanced metabolic activity. A significantly increased mitochondrial content in plasma treated cells supported this notion. On THP-1 cell proteome level, we identified an increase of protein translation with key regulatory proteins being involved in redox regulation (hypoxia inducible factor 2α), differentiation (retinoic acid signaling and interferon inducible factors), and cell growth (Yin Yang 1). Regulation of inflammation is a key element in many chronic diseases, and we found a significantly increased expression of the anti-inflammatory heme oxygenase 1 (HMOX1) and of the neutrophil attractant chemokine interleukin-8 (IL-8). Together, these results foster the view that cold physical plasma modulates the redox balance and inflammatory processes in wound related cells.
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    Sublethal injury and Viable but Non-culturable (VBNC) state in microorganisms during preservation of food and biological materials by non-thermal processes
    (Lausanne : Frontiers Media S. A, 2018) Schottroff, F.; Fröhling, A.; Zunabovic-Pichler, M.; Krottenthaler, A.; Schlüter, O.; Jäger, H.
    The viable but non-culturable (VBNC) state, as well as sublethal injury of microorganisms pose a distinct threat to food safety, as the use of traditional, culture-based microbiological analyses might lead to an underestimation or a misinterpretation of the product's microbial status and recovery phenomena of microorganisms may occur. For thermal treatments, a large amount of data and experience is available and processes are designed accordingly. In case of innovative inactivation treatments, however, there are still several open points with relevance for the investigation of inactivation mechanisms as well as for the application and validation of the preservation processes. Thus, this paper presents a comprehensive compilation of non-thermal preservation technologies, i.e., high hydrostatic pressure (HHP), pulsed electric fields (PEFs), pulsed light (PL), and ultraviolet (UV) radiation, as well as cold plasma (CP) treatments. The basic technological principles and the cellular and molecular mechanisms of action are described. Based on this, appropriate analytical methods are outlined, i.e., direct viable count, staining, and molecular biological methods, in order to enable the differentiation between viable and dead cells, as well as the possible occurrence of an intermediate state. Finally, further research needs are outlined.
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    Consensus model of a cyanobacterial light-dependent protochlorophyllide oxidoreductase in its pigment-free apo-form and photoactive ternary complex
    (London : Springer Nature, 2019) Schneidewind, Judith; Krause, Frank; Bocola, Marco; Stadler, Andreas Maximilian; Davari, Mehdi D.; Schwaneberg, Ulrich; Jaeger, Karl-Erich; Krauss, Ulrich
    Photosynthetic organisms employ two different enzymes for the reduction of the C17 = C18 double bond of protochlorophyllide (Pchlide), yielding the chlorophyll precursor chlorophyllide. First, a nitrogenase-like, light-independent (dark-operative) Pchlide oxidoreductase and secondly, a light-dependent Pchlide oxidoreductase (LPOR). For the latter enzyme, despite decades of research, no structural information is available. Here, we use protein structure modelling, molecular dynamics (MD) simulations combined with multi-wavelength analytical ultracentrifugation (MWA-AUC) and small angle X-ray scattering (SAXS) experiments to derive a consensus model of the LPOR apoprotein and the substrate/cofactor/LPOR ternary complex. MWA-AUC and SAXS experiments independently demonstrate that the apoprotein is monomeric, while ternary complex formation induces dimerization. SAXS-guided modelling studies provide a full-length model of the apoprotein and suggest a tentative mode of dimerization for the LPOR ternary complex, supported by published cross-link constraints. Our study provides a first impression of the LPOR structural organization.
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    Breast Cancer Stem Cell–Derived Tumors Escape from γδ T-cell Immunosurveillance In Vivo by Modulating γδ T-cell Ligands
    (Philadelphia, Pa. : AACR, 2023) Raute, Katrin; Strietz, Juliane; Parigiani, Maria Alejandra; Andrieux, Geoffroy; Thomas, Oliver S.; Kistner, Klaus M.; Zintchenko, Marina; Aichele, Peter; Hofmann, Maike; Zhou, Houjiang; Weber, Wilfried; Boerries, Melanie; Swamy, Mahima; Maurer, Jochen; Minguet, Susana
    There are no targeted therapies for patients with triple-negative breast cancer (TNBC). TNBC is enriched in breast cancer stem cells (BCSC), which play a key role in metastasis, chemoresistance, relapse, and mortality. γδ T cells hold great potential in immunotherapy against cancer and might provide an approach to therapeutically target TNBC. γδ T cells are commonly observed to infiltrate solid tumors and have an extensive repertoire of tumor-sensing mechanisms, recognizing stress-induced molecules and phosphoantigens (pAgs) on transformed cells. Herein, we show that patient-derived triple-negative BCSCs are efficiently recognized and killed by ex vivo expanded γδ T cells from healthy donors. Orthotopically xenografted BCSCs, however, were refractory to γ δ T-cell immunotherapy. We unraveled concerted differentiation and immune escape mechanisms: xenografted BCSCs lost stemness, expression of γ δ T-cell ligands, adhesion molecules, and pAgs, thereby evading immune recognition by γ δ T cells. Indeed, neither promigratory engineered γ δ T cells, nor anti–PD-1 checkpoint blockade, significantly prolonged overall survival of tumor-bearing mice. BCSC immune escape was independent of the immune pressure exerted by the γ δ T cells and could be pharmacologically reverted by zoledronate or IFNα treatment. These results pave the way for novel combinatorial immunotherapies for TNBC.