1,721,025 research outputs found
Interaction of Fusarium Mycotoxins, Fusaproliferin, and Fumonisin B1 with DNA Studied by Electrospray Ionization Mass Spectrometry
No full textInteraction of Fusarium Mycotoxins, Fusaproliferin, and Fumonisin B1 with DNA Studied by Electrospray Ionization Mass Spectrometry
No full text“Procedura di isolamento e di purificazione degli esosomi urinari per la ricerca di biomarcatori proteici”
No full textA multiplex quantitative proteomics strategy for protein biomarker studies in urinary exosomes
No full textUrinary exosomes have received considerable attention as a potential biomarker source for the diagnosis of renal diseases. Notwithstanding, their use in protein biomarker research is hampered by the lack of efficient methods for vesicle isolation, lysis, and protein quantification. Here we report an improved ultracentrifugation-based method that facilitates the solubilization and removal of major impurities associated with urinary exosomes. A double-cushion sucrose/D2O centrifugation step was used after a two-step differential centrifugation to separate exosomes from the heavier vesicles. After the removal of uromodulin, 378 and 79 unique proteins were identified, respectively, in low- and high-density fractions. Comparison of our data with two previously published data sets helped to define proteins commonly found in urinary exosomes. Lysis, protein extraction, and in-solution digestion of exosomes were then optimized for MudPIT application. More than a hundred exosomal proteins were quantified by four-plex iTRAQ analysis of single and pooled samples from two different age groups. For healthy men, six proteins (TSN1, PODXL, IDHC, PPAP, ACBP, and ANXA5) showed significant expression differences between exosome pools of those aged 25-50 and 50-70 years old. Thus, exosomes isolated by our method provide the basis for the development of robust quantitative methods for protein biomarker research
Reactivity of antineoplastic drugs with model peptides studied by advanced mass spectrometry methodologies
No full textThe human EV membranome
No full textExtracellular vesicles (EVs) are membrane bound structures that gained attention over the last decades because they mediate the horizontal transfer of their cargo molecules. Here, we have built the human EV membranome database by the combination of two in silico datasets obtained by: (i) GO term analysis of the human EV proteome and (ii) the extraction of proteins detected in EV preparations from the human membranome database. The EV membranome database contains 1299 frequently identified EV membrane proteins (MPs) and was used for enrichment analysis as well as for data mining of proteins belonging to relevant functional classes. We show that the majority of the EV MPs have plasma membrane origin. However, the presence of MPs of late-endosomal origin and the enrichment of exocytosis related GO term confirms the endosomal route as an alternative way to plasma membrane budding for the secretion of certain EV populations. Enrichment in proteins binding RNA, lipid and carbohydrate highlights the role of EV MPs as signals or ligands in intercellular communication. There are 18 tetraspanins and 40 glycophosphatidylinositol-anchored proteins in the EV membranome that have been frequently identified in EV isolates. Analysis of EV proteome revealed several G-protein-coupled receptor binding proteins (25 proteins) implicated in EV uptake and a high number of transmembrane receptor and tyrosine kinases signaling pathways related surface receptors (80 proteins) involved in cellular communication. These functional classes of EV MPs can represent important pharmaceuticals and clinical targets
Observation of Non-covalent Interactions Between Beauvericin and Oligonucleotides Using Electrospray Ionization Mass Spectrometry
No full textPhosphorylation of B14.5a subunit from bovine heart complex I identified by titanium dioxide selective enrichment and shotgun proteomics
No full textShotgun proteomics was used to study the steady phosphorylation state of NADH:ubiquinone oxidoreductase (complex I) subunits from bovine heart mitochondria. A total tryptic digestion of enzymatically active complex I was performed, and the resulting peptide mixture was subjected to phosphopeptide enrichment by the use of titanium dioxide (TiO2). The phosphopeptide-enriched fraction was separated and analyzed with nanoscale reverse-phase HPLC-ESI-MS/MS in single information-dependent acquisition. Hence two phosphorylated complex I subunits were detected: 42 kDa and B14.5a. Phosphorylation of 42-kDa subunit at Ser-59 has already been determined with fluorescent phosphoprotein-specific gel staining and mass spectrometry (Schilling, B., Aggeler, R., Schulenberg, B., Murray, J., Row, R. H., Capaldi, R. A., and Gibson, B. W. (2005) Mass spectrometric identification of novel phosphorylation site in subunit NDUFA10 of bovine mitochondrial complex I. FEBS Lett. 579, 2485-2490). In our work, this finding was confirmed using a non-gel-based approach. In addition, we report novel phosphorylation on B14.5a nuclear encoded subunit. We demonstrated evidence of the phosphorylation site at Ser-95 residue by collision-induced dissociation experiments on three different molecular ions of two tryptic phosphopeptides of B14.5a
Structural analysis of styrene oxide/haemoglobin adducts by mass spectrometry: identification of suitable biomarkers for human exposure evaluation
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