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1.
Quantitative assessment of methyl‐esterification and other side reactions in a standard propionylation protocol for detection of histone modifications
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Veerle Paternoster Anders Valdemar Edhager Christian Sibbersen Anders Lade Nielsen Anders Dupont Børglum Jane Hvarregaard Christensen Johan Palmfeldt 《Proteomics》2016,16(14):2059-2063
Histone modifications play an important role in regulating chromatin stability and gene expression, but to date, investigating them remains challenging. In order to obtain peptides suitable for MS‐based analysis, chemical derivatization of N‐terminus and lysine residues by propionic anhydride is commonly performed. Several side reactions (methyl‐esterification, amidation, solvolysis, overpropionylation, and missed propionylation) during propionylation protocols have been described, yet their relative abundances remain vague. Because methyl‐esterification could interfere with correct interpretation of the modification pattern, it is essential to take measures to avoid it. Here we present in‐depth quantitative analyses of methyl‐esterification and the other side reactions in a standard propionylation protocol containing methanol, and when replacing methanol with isopropanol or acetonitrile. We show that the use of alternative solvents can eliminate methyl‐esterification and that even though other side reactions are not prevented, their contribution can be kept relatively small. We also show that replacing methanol can be of importance also in other proteomics methods, such as mixed cation exchange, using methanol under acidic conditions. 相似文献
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Lavinia Paternoster David M. Evans Ellen Aagaard Nohr Claus Holst Valerie Gaborieau Paul Brennan Anette Prior Gjesing Niels Grarup Daniel R. Witte Torben J?rgensen Allan Linneberg Torsten Lauritzen Anelli Sandbaek Torben Hansen Oluf Pedersen Katherine S. Elliott John P. Kemp Beate St. Pourcain George McMahon Diana Zelenika J?rg Hager Mark Lathrop Nicholas J. Timpson George Davey Smith Thorkild I. A. S?rensen 《PloS one》2011,6(9)
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Lippolis R Gnoni A Abbrescia A Panelli D Maiorano S Paternoster MS Sardanelli AM Papa S Gaballo A 《Journal of Proteomics》2011,74(12):2846-2855
A comparative proteomic approach, using two dimensional gel electrophoresis and mass spectrometry, has been developed to compare and elucidate the differences among the cellular proteomes of four closely related isogenic O/C, SIN, N/R and T, B. clausii strains during both exponential and stationary phases of growth. Image analysis of the electropherograms reveals a high degree of concordance among the four proteomes, some proteins result, however, differently expressed. The proteins spots exhibiting high different expression level were identified, by mass-spectrometry analysis, as alcohol dehydrogenase (ADHA, EC1.2.1.3; ABC0046 isoform) aldehyde dehydrogenase (DHAS, EC 1.2.1.3; ABC0047 isoform) and flagellin-protein of B. clausii KSM-k16. The different expression levels of the two dehydrogenases were confirmed by quantitative RT-PCR and dehydrogenases enzymatic activity. The different patterns of protein expression can be considered as cell proteome signatures of the different strains. 相似文献
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Ambruosi B Uranio MF Sardanelli AM Pocar P Martino NA Paternoster MS Amati F Dell'Aquila ME 《PloS one》2011,6(11):e27452
Phthalates are ubiquitous environmental contaminants because of their use in plastics and other common consumer products. Di-(2-ethylhexyl) phthalate (DEHP) is the most abundant phthalate and it impairs fertility by acting as an endocrine disruptor. The aim of the present study was to analyze the effects of in vitro acute exposure to DEHP on oocyte maturation, energy and oxidative status in the horse, a large animal model. Cumulus cell (CC) apoptosis and oxidative status were also investigated. Cumulus-oocyte complexes from the ovaries of slaughtered mares were cultured in vitro in presence of 0.12, 12 and 1200 μM DEHP. After in vitro maturation (IVM), CCs were removed and evaluated for apoptosis (cytological assessment and TUNEL) and intracellular reactive oxygen species (ROS) levels. Oocytes were evaluated for nuclear chromatin configuration. Matured (Metaphase II stage; MII) oocytes were further evaluated for cytoplasmic energy and oxidative parameters. DEHP significantly inhibited oocyte maturation when added at low doses (0.12 μM; P<0.05). This effect was related to increased CC apoptosis (P<0.001) and reduced ROS levels (P<0.0001). At higher doses (12 and 1200 μM), DEHP induced apoptosis (P<0.0001) and ROS increase (P<0.0001) in CCs without affecting oocyte maturation. In DEHP-exposed MII oocytes, mitochondrial distribution patterns, apparent energy status (MitoTracker fluorescence intensity), intracellular ROS localization and levels, mt/ROS colocalization and total SOD activity did not vary, whereas increased ATP content (P<0.05), possibly of glycolytic origin, was found. Co-treatment with N-Acetyl-Cysteine reversed apoptosis and efficiently scavenged excessive ROS in DEHP-treated CCs without enhancing oocyte maturation. In conclusion, acute in vitro exposure to DEHP inhibits equine oocyte maturation without altering ooplasmic energy and oxidative stress parameters in matured oocytes which retain the potential to be fertilized and develop into embryos even though further studies are necessary to confirm this possibility. 相似文献
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Background
Whether inhaled corticosteroids suppress airway inflammation in chronic obstructive pulmonary disease (COPD) remains controversial. We sought to determine the effects of inhaled corticosteroids on sputum indices of inflammation in stable COPD.Methods
We searched MEDLINE, EMBASE, CINAHL, and the Cochrane Databases for randomized, controlled clinical trials that used induced sputum to evaluate the effect of inhaled corticosteroids in stable COPD. For each chosen study, we calculated the mean differences in the concentrations of sputum cells before and after treatment in both intervention and control groups. These values were then converted into standardized mean differences to accommodate the differences in patient selection, clinical treatment, and biochemical procedures that were employed across original studies. If significant heterogeneity was present (p < 0.10), then a random effects model was used to pool the original data. In the absence of significant heterogeneity, a fixed effects model was used.Results
We identified six original studies that met the inclusion criteria (N = 162 participants). In studies with higher cumulative dose (≥ 60 mg) or longer duration of therapy (≥ 6 weeks), inhaled corticosteroids were uniformly effective in reducing the total cell, neutrophil, and lymphocyte counts. In contrast, studies with lower cumulative dose (< 60 mg) or shorter duration of therapy (< 6 weeks) did not demonstrate a favorable effect of inhaled corticosteroids on these sputum indices.Conclusions
Our study suggests that prolonged therapy with inhaled corticosteroids is effective in reducing airway inflammation in stable COPD. 相似文献7.
Significance of nucleotide sequence alignments: a method for random sequence permutation that preserves dinucleotide and codon usage 总被引:10,自引:0,他引:10
The similarity of two nucleotide sequences is often expressed in terms of
evolutionary distance, a measure of the amount of change needed to
transform one sequence into the other. Given two sequences with a small
distance between them, can their similarity be explained by their base
composition alone? The nucleotide order of these sequences contributes to
their similarity if the distance is much smaller than their average
permutation distance, which is obtained by calculating the distances for
many random permutations of these sequences. To determine whether their
similarity can be explained by their dinucleotide and codon usage, random
sequences must be chosen from the set of permuted sequences that preserve
dinucleotide and codon usage. The problem of choosing random dinucleotide
and codon-preserving permutations can be expressed in the language of graph
theory as the problem of generating random Eulerian walks on a directed
multigraph. An efficient algorithm for generating such walks is described.
This algorithm can be used to choose random sequence permutations that
preserve (1) dinucleotide usage, (2) dinucleotide and trinucleotide usage,
or (3) dinucleotide and codon usage. For example, the similarity of two
60-nucleotide DNA segments from the human beta-1 interferon gene
(nucleotides 196-255 and 499-558) is not just the result of their nonrandom
dinucleotide and codon usage.
相似文献
8.
Litzow MR Dietz AB Bulur PA Butler GW Gastineau DA Hoering A Fink SR Letendre L Padley DJ Paternoster SF Tefferi A Vuk-Pavlović S 《Cytotherapy》2006,8(3):290-298
BACKGROUND: We conducted a phase I clinical immunotherapy trial of CML to evaluate the safety of a clinical-grade leukemic DC product standardized for purity and mature phenotype. METHODS: We injected autologous DC into patients in late chronic or accelerated phases of CML. The patients received mature CD83+ and bcr-abl+ DC prepared from CD14+ cells. Two cohorts of three patients received four injections each of 3 x 10(6) DC and 15 x 10(6) DC/injection, respectively. The first patient was studied before imatinib mesylate (IM) was available, four patients were treated concurrently with IM therapy and one did not tolerate the IM and was off the drug at the time of DC therapy. IM effects on WBC counts precluded DC preparation in numbers sufficient for further dose escalation. The first patient received DC s.c. and all subsequent patients received DC into a cervical lymph node under ultrasound guidance. RESULTS: DC injections were well tolerated. We observed no clinical responses. T cells drawn later in the course of therapy were more sensitive to stimulation by CML DC in vitro. DISCUSSION: The increase in T-cell sensitivity to CML-specific stimulation that accompanied active immunization by CML DC justifies further clinical studies, possibly with modifications such as an increased frequency and number of DC injections. 相似文献
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Benthem L Keizer K Wiegman CH de Boer SF Strubbe JH Steffens AB Kuipers F Scheurink AJ 《American journal of physiology. Endocrinology and metabolism》2000,279(6):E1286-E1293
We tested the hypothesis that excessive portal venous supply of long-chain fatty acids to the liver contributes to the development of insulin resistance via activation of the hypothalamus-pituitary-adrenal axis (HPA axis) and sympathetic system. Rats received an intraportal infusion of the long-chain fatty acid oleate (150 nmol/min, 24 h), the medium-chain fatty acid caprylate, or the solvent. Corticosterone (Cort) and norepinephrine (NE) were measured as indexes for HPA axis and sympathetic activity, respectively. Insulin sensitivity was assessed by means of an intravenous glucose tolerance test (IVGTT). Oleate infusion induced increases in plasma Cort (Delta = 13.5 +/- 3.6 microg/dl; P < 0.05) and NE (Delta = 235 +/- 76 ng/l; P < 0.05), whereas caprylate and solvent had no effect. The area under the insulin response curve to the IVGTT was larger in the oleate-treated group than in the caprylate and solvent groups (area = 220 +/- 35 vs. 112 +/- 13 and 106 +/- 8, respectively, P < 0.05). The area under the glucose response curves was comparable [area = 121 +/- 13 (oleate) vs. 135 +/- 20 (caprylate) and 96 +/- 11 (solvent)]. The results are consistent with the concept that increased portal free fatty acid is involved in the induction of visceral obesity-related insulin resistance via activation of the HPA axis and sympathetic system. 相似文献
10.
Alex?SF?Doney Bettina?Fischer Simon?P?Lee Andrew?D?Morris Graham?Leese Colin?NA?PalmerEmail author 《Nuclear receptor》2005,3(1):4