Haptoglobin binding to apolipoprotein A-I prevents damage from hydroxyl radicals on its stimulatory activity of the enzyme lecithin-cholesterol acyl-transferase

Apolipoprotein A-I (ApoA-I), a major component of HDL, binds Haptoglobin, a plasma protein transporting to liver or macrophages free Hb for preventing hydroxyl radical production. This work aimed to assess whether Haptoglobin protects ApoA-I against this radical. Human ApoA-I structure, as analyzed by electrophoresis and MS, was found severely altered by hydroxyl radicals in vitro. Lower alteration of ApoA-I was found when HDL was oxidized in the presence of Haptoglobin. ApoA-I oxidation was limited also when the complex of Haptoglobin with both high density lipoprotein and Hb, immobilized on resin beads, was exposed to hydroxyl radicals. ApoA-I function to stimulate cholesterol esterification was assayed in vitro by using ApoA-I-containing liposomes. Decreased stimulation was observed when liposomes oxidized without Haptoglobin were used. Conversely, after oxidative stress in presence of Haptoglobin (0.5 microM monomer), the liposome activity did not change. Plasma of Carrageenan-treated mice was analyzed by ELISA for the levels of Haptoglobin and ApoA-I, and used to isolate HDL for MS analysis. Hydroxyproline-containing fragments of ApoA-I were found associated with low levels of Haptoglobin (18 microM monomer), whereas they were not detected when the Haptoglobin level increased (34-70 microM monomer). Therefore Haptoglobin, when circulating at enhanced levels with free Hb during the acute phase of inflammation, might protect ApoA-I structure and function against hydroxyl radicals

Covering Chemical Diversity of Genetically-Modified Tomatoes Using Metabolomics for Objective Substantial Equivalence Assessment

As metabolomics can provide a biochemical snapshot of an organism's phenotype it is a promising approach for charting the unintended effects of genetic modification. A critical obstacle for this application is the inherently limited metabolomic coverage of any single analytical platform. We propose using multiple analytical platforms for the direct acquisition of an interpretable data set of estimable chemical diversity. As an example, we report an application of our multi-platform approach that assesses the substantial equivalence of tomatoes over-expressing the taste-modifying protein miraculin. In combination, the chosen platforms detected compounds that represent 86% of the estimated chemical diversity of the metabolites listed in the LycoCyc database. Following a proof-of-safety approach, we show that % had an acceptable range of variation while simultaneously indicating a reproducible transformation-related metabolic signature. We conclude that multi-platform metabolomics is an approach that is both sensitive and robust and that it constitutes a good starting point for characterizing genetically modified organisms

Proteome analysis of biopsies from patients suffering from rectum carcinoma.

Napoli, 26-27 settembre 200

Uso dell'aptoglobina, peptidi leganti l'aptoglobina, polimeri contenenti gli stessi e loro uso

Sono stati sintetizzati peptidi mimetici del dominio dell'apolipoproteina E (ApoE), che è legato dall'aptoglobina (Hpt). Questi peptidi possono essere utilizzati per fini diagnostici, per titolare Hpt come marker della fase acuta dell'infiammazione. Poiché i peptidi spiazzano ApoE da Hpt, essi possono anche essere usati per liberare ApoE da complessi con Hpt e permetterne, quindi, la massima attività di stimolazione sull'attività di LCAT (un enzima che svolge un ruolo chiave nel trasporto inverso del colesterolo e, quindi, nella prevenzione di malattie cardiovascolari)

Differential protein expression in Parkinson\u2019s Disease

Napoli, 26-27 settembre 2003

Proteomic analysis of a genetically modified maize flour carrying the CRY1AB gene and comparison to the corresponding wild-type

Protein expression in a maize hybrid flour (hereafter called WT) and its corresponding transgenic version resistant to European corn borer (hereafter called BT and carrying a gene encoding for the Bacillus thuringiensis insecticidal protein Cry1Ab) has been studied by means of two-dimensional gel electrophoresis coupled with mass spectrometry. This comparison has been chosen as a model to verify proteomics capability in detecting unexpected differences between near-isogenic lines (differing in the ideal case only for the presence of the transgenic protein). Some unpredictable differences were detected: i) glucose and ribitol dehydrogenase spot was unique of BT maize; ii) endochitinase A spot was unique of WT maize; iii) triosephosphate isomerase 1 and one spot of globulin-1 S were overexpressed while cytosolic 3-phosphoglycerate kinase and one spot of aldose reductase were down-regulated in BT maize with respect to WT. These results outline the potential of the new nontargeted \u201c\u2013omics\u201d technologies (in particular proteomics) in the detection of unexpected, unintended and unwanted variations in Genetically Modified (GM) versus non-GM food comparison and suggest the possible employment of these technologies in substantial equivalence evaluation. Moreover, a reference map for maize flour was built: forty spots, corresponding to twenty-five different proteins, were successfully identified

Proteomics as a tool to improve investigation of substantial equivalence in genetically modified organisms: The case of a virus-resistant tomato

At present, the so-called "substantial equivalence" is the only widely accepted criterion for deciding whether or not a transgenic food is, from an alimentary point of view, to be considered totally correspondent to the "traditional" one from which it derives. Although never exactly defined, it deals with a comparison between the chemical composition of the two foods. A more in-depth analysis can be performed by one of the most suitable methods that allows for the simultaneous screening of many components without prior identification, the analysis of the proteome. As a model for testing this kind of approach, we compared protein expression of two types of tomato plants, having the same genetic background, except for a virus resistance trait introduced by genetic engineering. When proteins extracted from seedlings of the two types were analyzed by two-dimensional electrophoresis, no significant differences, either qualitative or quantitative, were detected, indicating that in this case the expression of major proteins was unmodified by the genetic manipulation. Fifteen proteins were identified by peptide mass fingerprinting