Search results for " High pressure liquid"
showing 10 items of 705 documents
PLA2-mediated catalytic activation of its inhibitor 25-acetyl-petrosaspongiolide M: serendipitous identification of a new PLA2 suicide inhibitor.
2004
Abstract25-Acetyl-petrosaspongiolide M (PMAc) (1), a mild non-covalent PLA2 inhibitor, unexpectedly recovers, after incubation with bvPLA2, the ability to covalently modify the enzyme target. This study demonstrates the catalytic effect of bvPLA2 in converting 1 in its deacetylated congener petrosaspongiolide M (PM) (2), a strong covalent PLA2 inhibitor whose molecular mechanism of inhibition has already been clarified. Moreover, our findings outline the potential role of PMAc as anti-inflammatory pro-drug, by virtue of its ability of delivering the active PM agent at the site of inflammation, functioning as a suicide inhibitor.
Molecular mass of macromolecules and subunits and the quaternary structure of hemoglobin from the microcrustacean Daphnia magna
2006
The molecular masses of macromolecules and subunits of the extracellular hemoglobin from the fresh-water crustacean Daphnia magna were determined by analytical ultracentrifugation, multiangle laser light scattering and electrospray ionization mass spectrometry. The hemoglobins from hypoxia-incubated, hemoglobin-rich and normoxia-incubated, hemoglobin-poor Daphnia magna were analyzed separately. The sedimentation coefficient of the macromolecule was 17.4 +/- 0.1 S, and its molecular mass was 583 kDa (hemoglobin-rich animals) determined by AUC and 590.4 +/- 11.1 kDa (hemoglobin-rich animals) and 597.5 +/- 49 kDa (hemoglobin-poor animals), respectively, determined by multiangle laser light sca…
Expression and glycosylation studies of human FGF receptor 4
2001
Fibroblast growth factor receptor subtype 4 (FGFR4) has been shown to have special activation properties and just one splicing form, unlike the other FGFRs. FGFR4 overexpression is correlated with breast cancer and therefore FGFR4 is a target for drug design. Our aim is to overexpress high amounts of homogeneous FGFR4 extracellular domain (FGFR4ed) for structural studies. We show that baculovirus-insect cell-expressed FGFR4ed is glycosylated on three (N88, N234, and N266) of the six possible N-glycosylation sites but is not O-glycosylated. The deglycosylated triple mutant was expressed and had binding properties similar to those of glycosylated FGFR4ed, but was still heterogeneous. Large am…
Evolution of nacre: biochemistry and proteomics of the shell organic matrix of the cephalopod Nautilus macromphalus.
2009
12 pages; International audience; In mollusks, one of the most widely studied shell textures is nacre, the lustrous aragonitic layer that constitutes the internal components of the shells of several bivalves, a few gastropods, and one cephalopod: the nautilus. Nacre contains a minor organic fraction, which displays a wide range of functions in relation to the biomineralization process. Here, we have biochemically characterized the nacre matrix of the cephalopod Nautilus macromphalus. The acid-soluble matrix contains a mixture of polydisperse and discrete proteins and glycoproteins, which interact with the formation of calcite crystals. In addition, a few bind calcium ions. Furthermore, we h…
Identification of Prostate-Enriched Proteins by In-depth Proteomic Analyses of Expressed Prostatic Secretions in Urine
2012
Urinary expressed prostatic secretion or "EPS-urine" is proximal tissue fluid that is collected after a digital rectal exam (DRE). EPS-urine is a rich source of prostate-derived proteins that can be used for biomarker discovery for prostate cancer (PCa) and other prostatic diseases. We previously conducted a comprehensive proteome analysis of direct expressed prostatic secretions (EPS). In the current study, we defined the proteome of EPS-urine employing Multidimensional Protein Identification Technology (MudPIT) and providing a comprehensive catalogue of this body fluid for future biomarker studies. We identified 1022 unique proteins in a heterogeneous cohort of 11 EPS-urines derived from …
Metaproteomic analysis of atmospheric aerosol samples.
2016
Metaproteomic analysis of air particulate matter provides information about the abundance and properties of bioaerosols in the atmosphere and their influence on climate and public health. We developed and applied efficient methods for the extraction and analysis of proteins from glass fiber filter samples of total, coarse, and fine particulate matter. Size exclusion chromatography was applied to remove matrix components, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied for protein fractionation according to molecular size, followed by in-gel digestion and LC-MS/MS analysis of peptides using a hybrid Quadrupole-Orbitrap MS. Maxquant software and the Swiss-…
Enhancing Sensitivity of Microflow-Based Bottom-Up Proteomics through Postcolumn Solvent Addition.
2019
The introduction of more sensitive mass spectrometers allows researchers to adapt front-end liquid chromatography (LC) to individual needs for the analysis of complex proteomes. Where absolute sensitivity is not paramount, it is advantageous to switch from a highly sensitive nanoflow-LC setup, the de facto standard platform in mass-spectrometry (MS)-based discovery proteomics, to a more robust, high-throughput-compatible microflow or conventional-flow setup. To enhance the microflow-LC-MS electrospray process of complex proteomic samples, we tested the effects of different solvents, including 2-propanol, methanol, and acetonitrile, pure or as mixture with dimethyl sulfoxide, which were adde…
Ultrahigh-Throughput Proteomics Using Fast RPLC Separations with ESI-MS/MS
2005
We describe approaches for proteomics analysis using electrospray ionization-tandem mass spectrometry coupled with fast reversed-phase liquid chromatography (RPLC) separations. The RPLC separations used 50-microm-i.d. fused-silica capillaries packed with submicrometer-sized C18-bonded porous silica particles and achieved peak capacities of 130-420 for analytes from proteome tryptic digests. When these separations were combined with linear ion trap tandem mass spectrometry measurements, approximately 1000 proteins could be identified in 50 min from approximately 4000 identified tryptic peptides; approximately 550 proteins in 20 min from approximately 1800 peptides; and approximately 250 prot…
Column switching and high-performance liquid chromatography in the analysis of amitriptyline, nortriptyline and hydroxylated metabolites in human pla…
1992
A column-switching system for the direct injection of plasma or serum samples, followed by isocratic high-performance liquid chromatography and ultraviolet detection, is described for the simultaneous quantitation of the tricyclic antidepressant amitriptyline, its demethylated metabolite nortriptyline and the E- and Z-isomers of 10-hydroxyamitriptyline and 10-hydroxynortriptyline. The method included adsorption of amitriptyline and metabolites on a reversed-phase C8 clean-up column (10 microns; 20 mm x 4.6 mm I.D.), washing of unwanted material to waste and, after on-line column-switching, separation on a cyanopropyl analytical column (5 microns; 250 mm x 4.6 mm I.D.). The compounds of inte…
Differential changes in purine nucleotides after Doxorubicin treatment of human cancer cells in vitro
2002
The present investigation was performed to elucidate the role of purine nucleotides as potential indicators of chemosensitivity of malignant tumors. Drug-sensitive (s) and -resistant (r) tumor cell lines grown as monolayers (s: T47D, MCF-7 wild-type; r: NCI/ADR-RES, MCF-7/MDR) or as multicellular spheroids (T47D; NCI/ADR-RES) were exposed to 0.1, 1.0, and 10.0 microM Doxorubicin for up to 24 h. Purine nucleotides were assayed using HPLC and with some selected spheroids using imaging bioluminescence. The data show that in the time frame of the experiments reproducible and statistically significant changes in the nucleotides only occur at the highest drug concentration investigated. Under the…