Project description:Identification of targets of the protein disulfide reductase thioredoxin using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) and thiol specific differential labeling with isotope-coded affinity tags (ICAT). Reduction of specific target disulfides is quantified by measuring ratios of cysteine residues labeled with the heavy (13C) and light (12C) ICAT reagents in peptides derived from tryptic digests of Trx-treated and non-treated samples. Keywords: protein, LC-MS/MS, ICAT
Project description:This dataset is part of a study aimed at developing algorithms for the quantification of stable isotope content in microorganisms after labeling them with stable isotope-labeled substrates. In this dataset Escherichia coli cultures were labeled with different percentages (1% or 10%) of either single-carbon 13C glucose (13C2) or fully-labeled 13C glucose (13C1-6). Labeled cells were subsequently mixed with unlabeled E. coli cells in fixed ratios (50%, 90%, 95%, 99%). Cultures of E. coli were grown in M9 minimal medium in which a percentage of the glucose was replaced with 13C2 or 13C1-6 glucose for >10 generations to achieve close to complete labeling of cells. Triplicate cultures were grown for each percentage. Please note that the unlabeled glucose that was used of course had a natural content of 13C of around 1.1%, thus the 0% added label samples have an actual 13C content of 1.1% and all added label is on top of this value. We included a tab delimited table with this submission providing details on all raw files.
Project description:This dataset is part of a study aimed at developing algorithms for the quantification of stable isotope content in microorganisms after labeling them with stable isotope-labeled substrates. In this dataset Escherichia coli and Bacillus subtilis cultures were labeled with different percentages of single-carbon 13C glucose (13C2). Cultures of B. subtilis and E. coli were grown in Bacillus minimal medium or M9 minimal medium (E. coli) in which a percentage of the glucose was replaced with 13C2 glucose for >10 generations to achieve close to complete labeling of cells. The following percentages of 13C2 glucose were added 0, 0.01, 0.025, 0.1, 0.25, 1, 5 and 10%. Triplicate cultures were grown for each percentage. Please note that the unlabeled glucose that was used of course had a natural content of 13C of around 1.1%, thus the 0% added label samples have an actual 13C content of 1.1% and all added label is on top of this value. We included a tab delimited table with this submission providing details on all raw files.
Project description:This dataset is part of a study aimed at developing algorithms for the quantification of stable isotope content in microorganisms after labeling them with stable isotope-labeled substrates. In this dataset Escherichia coli and Bacillus subtilis cultures were labeled with different percentages of fully labeled 13C glucose (13C6). Cultures of B. subtilis and E. coli were grown in Bacillus minimal medium or M9 minimal medium (E. coli) in which a percentage of the glucose was replaced with 13C6 glucose for >10 generations to achieve close to complete labeling of cells. The following percentages of 13C6 glucose were added 0, 0.01, 0.025, 0.1, 0.25, 1, 5 and 10%. Triplicate cultures were grown for each percentage. Please note that the unlabeled glucose that was used of course had a natural content of 13C of around 1.1%, thus the 0% added label samples have an actual 13C content of 1.1% and all added label is on top of this value. We included a tab delimited table with this submission providing details on all raw files.
2022-12-30 | PXD023693 | Pride
Project description:Effect of 13C-labled straw on the results of DNA Stable isotope probing experiments
Project description:This dataset is part of a study aimed at developing algorithms for the quantification of stable isotope content in microorganisms in microbial communities after labeling them with stable isotope-labeled substrates. For this dataset Escherichia coli cultures were labeled with different percentages (1, 5 and 10%) of fully labeled 13C glucose (13C1-6) and spiked-in into a mock microbial community consisting of 32 species of bacteria, archaea, eukaryote and bacteriophages (UNEVEN Community described in Kleiner et al. 2017 Nat Communications 8(1):1558). The community also contained unlabeled E. coli cells and labeled/unlabeled E. coli cells in the spike-in sample were at a 1:1 ratio. Cultures of E. coli were grown in M9 minimal medium in which a percentage of the glucose was replaced with 13C1-6 glucose for >10 generations to achieve close to complete labeling of cells. The following percentages of 13C1-6 glucose were added 1, 5 and 10%. Triplicate cultures were grown for each percentage. Please note that the unlabeled glucose that was used of course had a natural content of 13C of around 1.1%, thus the 0% added label samples have an actual 13C content of 1.1% and all added label is on top of this value. We included a tab delimited table with this submission providing details on all raw files.
2022-12-30 | PXD024174 | Pride
Project description:Identification of acetaminophen assimilating microorganisms in mixed microbial communities using 13C-DNA stable isotope probing
Project description:Chemosynthetic symbioses occur worldwide in marine habitats, but comprehensive physiological studies of chemoautotrophic bacteria thriving on animals are scarce. Stilbonematinae are coated by monocultures of thiotrophic Gammaproteobacteria. As these nematodes migrate through the redox zone, their ectosymbionts experience varying oxygen concentrations. Here, by applying omics, Raman microspectroscopy and stable isotope labeling, we investigated the effect of oxygen on the metabolism of Candidatus Thiosymbion oneisti. Unexpectedly, sulfur oxidation genes were upregulated in anoxic relative to oxic conditions, but carbon fixation genes and incorporation of 13C-labeled bicarbonate were not. Instead, several genes involved in carbon fixation, organic carbon assimilation and polyhydroxyalkanoate (PHA) biosynthesis, as well as nitrogen fixation and urea utilization were upregulated in oxic conditions. Furthermore, in the presence of oxygen, stress-related genes were upregulated together with vitamin biosynthesis genes likely necessary to withstand its deleterious effects, and fewer symbionts were detected to divide. Based on this first global physiological study of an uncultured chemosynthetic ectosymbiont, we propose that, in anoxic sediment, its proliferation is powered by anaerobic sulfur oxidation coupled to denitrification, whereas in upper layers it makes use of aerobic respiration to facilitate assimilation of carbon and nitrogen, and to survive oxidative stress. The ectosymbiont’s versatile metabolism is thus well-adapted to exploiting a highly changeable environment.
Project description:Obesity is a major risk factor for the development of insulin resistance and type II diabetes. The nuclear receptors PPAR delta and PPAR gamma play a central role in regulating metabolism in adipose tissue, as well as being targets for the treatment of insulin resistance. The metabolic effects of PPAR delta and PPAR gamma activation have been examined both in vivo in white adipose tissue from ob/ob mice and in vitro in cultured 3T3-L1 adipocytes using a combined 1H NMR spectroscopy and mass spectrometry metabolomic methodology to understand the contrasting roles of these receptors. These steady state measurements were supplemented with 13C-stable isotope substrate labeling to assess fluxes, respirometry and transcriptomic microarray analysis. The metabolic effects of the two receptors were readily distinguished, with PPAR gamma ?activation characterised by increased fat storage and fat synthesis/elongation, while activation of PPAR delta caused increased fatty acid beta-oxidation, TCA cycle rate and oxidation of extracellular branch chain amino acids. Stimulated glycolysis and increased desaturation of fatty acids were the only common pathways. PPAR delta has a role as an anti-obesity target as well as an anti-diabetic. Total RNA obtained from cultured 3T3-L1 cells treated for 48 hours with either DMSO control, GW610742 PPARd agonist or GW347845 PPARg agonist and compared.