Publications by authors named "Manuel José Rodríguez-Ortega"

2 Publications

  • Page 1 of 1

Proteomics characterization of outer membrane vesicles from the extraintestinal pathogenic Escherichia coli DeltatolR IHE3034 mutant.

Mol Cell Proteomics 2008 Mar 2;7(3):473-85. Epub 2007 Nov 2.

Biochemistry and Molecular Biology Unit, Novartis Vaccines and Diagnostics, 53100 Siena, Italy.

Extraintestinal pathogenic Escherichia coli are the cause of a diverse spectrum of invasive infections in humans and animals, leading to urinary tract infections, meningitis, or septicemia. In this study, we focused our attention on the identification of the outer membrane proteins of the pathogen in consideration of their important biological role and of their use as potential targets for prophylactic and therapeutic interventions. To this aim, we generated a DeltatolR mutant of the pathogenic IHE3034 strain that spontaneously released a large quantity of outer membrane vesicles in the culture supernatant. The vesicles were analyzed by two-dimensional electrophoresis coupled to mass spectrometry. The analysis led to the identification of 100 proteins, most of which are localized to the outer membrane and periplasmic compartments. Interestingly based on the genome sequences available in the current public database, seven of the identified proteins appear to be specific for pathogenic E. coli and enteric bacteria and therefore are potential targets for vaccine and drug development. Finally we demonstrated that the cytolethal distending toxin, a toxin exclusively produced by pathogenic bacteria, is released in association with the vesicles, supporting the recently proposed role of bacterial vesicles in toxin delivery to host cells. Overall, our data demonstrated that outer membrane vesicles represent an ideal tool to study Gram-negative periplasm and outer membrane compartments and to shed light on new mechanisms of bacterial pathogenesis.
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http://dx.doi.org/10.1074/mcp.M700295-MCP200DOI Listing
March 2008

Mutagenic activation of arylamines by subcellular fractions of Chamaelea gallina clams exposed to environmental pollutants.

Environ Mol Mutagen 2003 ;41(1):55-63

Department of Biochemistry and Molecular Biology, University of Córdoba, Campus de Rabanales, Severo Ochoa Building, 2nd Floor, 14071 Córdoba, Spain.

Biochemical measurements in the sentinel clam Chamaelea gallina have been used as biomarkers of marine pollution. In this study, S9, cytosolic fractions (CF), and microsomal fractions (MF) prepared from unexposed clams and clams exposed to model pollutants were used to activate 2-aminoanthracene (2-AA) and 2-acetylaminofluorene (AAF) to mutagens in Salmonella typhimurium strain BA149, which overexpresses O-acetyltransferase. Arylamine activation was similar with subcellular fractions from unexposed and Aroclor 1254-exposed clams, but decreased with fractions from As(III)- and Cu(II)-exposed clams. Bioactivation of arylamines by CF was higher than by MF, and higher with NADH than with NADPH as the reducing agent. alpha-Naphthoflavone inhibited AAF activation by CF and MF, but increased 2-AA activation nearly twofold. In contrast to the results with arylamine activation, benzo[a]pyrene hydroxylase (BPH) activity increased twofold in fractions from Aroclor 1254- and Cu(II)-exposed clams. Activation of 2-AA was also evaluated using S9 fractions from clams sampled at littoral sites with different pollutant levels. Compared to a reference site, lower 2-AA bioactivation and higher BPH activity were found in clams containing high levels of copper and organic contaminants, although the differences were not statistically significant. While these findings agree with the results of the model Cu(II) exposure, the effects of other pollutants cannot be ruled out. The results of the study demonstrate that arylamine activation by clams is not preferentially catalyzed by microsomal monooxygenases but by unknown cytosolic system(s), and that bioactivation of 2-AA and AAF appears to occur by different pathways.
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http://dx.doi.org/10.1002/em.10130DOI Listing
March 2003