Biomedical subjects

Ana Conesa

Publications and source records attributed to Ana Conesa.

7 recordsLinked to original sources

PubMed2006

Development of the GENIPOL European flounder (Platichthys flesus) microarray and determination of temporal transcriptional responses to cadmium at low dose.

We have constructed a high density, 13 270-clone cDNA array for the sentinel fish species European flounder (Platichthys flesus), combining clones from suppressive subtractive hybridization and a liver cDNA library; DNA sequences of 5211 clones were determined. Fish were treated by single intraperitoneal injection with 50 micrograms cadmium chloride per kilogram body weight, a dose relevant to environmental exposures, and hepatic gene expression changes were determined at 1, 2, 4, 8, and 16 days postinjection in comparison to saline-treated controls. Gene expression responses were confirmed by real-time reverse transcription polymerase chain reaction (RT-PCR). Blast2GO gene ontology analysis highlighted a general induction of the unfolded protein response, response to oxidative stress, protein synthesis, transport, and degradation pathways, while apoptosis, cell cycle, cytoskeleton, and cytokine genes were also affected. Transcript levels of cytochrome P450 1A (CYP1A) were repressed and vitellogenin altered, real-time PCR showed induction of metallothionein. We thus describe the establishment of a useful resource for ecotoxicogenomics and the determination of the temporal molecular responses to cadmium, a prototypical heavy metal pollutant.

Animals↗

PubMed2006

maSigPro: a method to identify significantly differential expression profiles in time-course microarray experiments.

MOTIVATION: Multi-series time-course microarray experiments are useful approaches for exploring biological processes. In this type of experiments, the researcher is frequently interested in studying gene expression changes along time and in evaluating trend differences between the various experimental groups. The large amount of data, multiplicity of experimental conditions and the dynamic nature of the experiments poses great challenges to data analysis. RESULTS: In this work, we propose a statistical procedure to identify genes that show different gene expression profiles across analytical groups in time-course experiments. The method is a two-regression step approach where the experimental groups are identified by dummy variables. The procedure first adjusts a global regression model with all the defined variables to identify differentially expressed genes, and in second a variable selection strategy is applied to study differences between groups and to find statistically significant different profiles. The methodology is illustrated on both a real and a simulated microarray dataset.

Algorithms↗

PubMed2005

Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research.

SUMMARY: We present here Blast2GO (B2G), a research tool designed with the main purpose of enabling Gene Ontology (GO) based data mining on sequence data for which no GO annotation is yet available. B2G joints in one application GO annotation based on similarity searches with statistical analysis and highlighted visualization on directed acyclic graphs. This tool offers a suitable platform for functional genomics research in non-model species. B2G is an intuitive and interactive desktop application that allows monitoring and comprehension of the whole annotation and analysis process. AVAILABILITY: Blast2GO is freely available via Java Web Start at http://www.blast2go.de. SUPPLEMENTARY MATERIAL: http://www.blast2go.de -> Evaluation.

Algorithms↗

PubMed2003

Examining the role of glutamic acid 183 in chloroperoxidase catalysis.

Site-directed mutagenesis has been used to investigate the role of glutamic acid 183 in chloroperoxidase catalysis. Based on the x-ray crystallographic structure of chloroperoxidase, Glu-183 is postulated to function on distal side of the heme prosthetic group as an acid-base catalyst in facilitating the reaction between the peroxidase and hydrogen peroxide with the formation of Compound I. In contrast, the other members of the heme peroxidase family use a histidine residue in this role. Plasmids have now been constructed in which the codon for Glu-183 is replaced with a histidine codon. The mutant recombinant gene has been expressed in Aspergillus niger. An analysis of the produced mutant gene shows that the substitution of Glu-183 with a His residue is detrimental to the chlorination and dismutation activity of chloroperoxidase. The activity is reduced by 85 and 50% of wild type activity, respectively. However, quite unexpectedly, the epoxidation activity of the mutant enzyme is significantly enhanced approximately 2.5-fold. These results show that Glu-183 is important but not essential for the chlorination activity of chloroperoxidase. It is possible that the increased epoxidation of the mutant enzyme is based on an increase in the hydrophobicity of the active site.

Aspergillus niger↗

PubMed2002

Fungal peroxidases: molecular aspects and applications.

Peroxidases are oxidoreductases that utilize hydrogen peroxide to catalyze oxidative reactions. A large number of peroxidases have been identified in fungal species and are being characterized at the molecular level. In this manuscript we review the current knowledge on the molecular aspects of this type of enzymes. We present an overview of the research efforts undertaken in deciphering the structural basis of the catalytic properties of fungal peroxidases and discuss molecular genetics and protein homology aspects of this enzyme class. Finally, we summarize the potential biotechnological applications of these enzymes and evaluate recent advances on their expression in heterologous systems for production purposes.

Amino Acid Sequence↗

PubMed2002

Filamentous fungi as cell factories for heterologous protein production.

Filamentous fungi have been used as sources of metabolites and enzymes for centuries. For about two decades, molecular genetic tools have enabled us to use these organisms to express extra copies of both endogenous and exogenous genes. This review of current practice reveals that molecular tools have enabled several new developments. But it has been process development that has driven the final breakthrough to achieving commercially relevant quantities of protein. Recent research into gene expression in filamentous fungi has explored their wealth of genetic diversity with a view to exploiting them as expression hosts and as a source of new genes. Inevitably, the progress in the 'genomics' technology will further develop high-throughput technologies for these organisms.

Fermentation↗

PubMed2002

Calnexin overexpression increases manganese peroxidase production in Aspergillus niger.

Heme-containing peroxidases from white rot basidiomycetes, in contrast to most proteins of fungal origin, are poorly produced in industrial filamentous fungal strains. Factors limiting peroxidase production are believed to operate at the posttranslational level. In particular, insufficient availability of the prosthetic group which is required for peroxidase biosynthesis has been proposed to be an important bottleneck. In this work, we analyzed the role of two components of the secretion pathway, the chaperones calnexin and binding protein (BiP), in the production of a fungal peroxidase. Expression of the Phanerochaete chrysosporium manganese peroxidase (MnP) in Aspergillus niger resulted in an increase in the expression level of the clxA and bipA genes. In a heme-supplemented medium, where MnP was shown to be overproduced to higher levels, induction of clxA and bipA was also higher. Overexpression of these two chaperones in an MnP-producing strain was analyzed for its effect on MnP production. Whereas bipA overexpression seriously reduced MnP production, overexpression of calnexin resulted in a four- to fivefold increase in the extracellular MnP levels. However, when additional heme was provided in the culture medium, calnexin overexpression had no synergistic effect on MnP production. The possible function of these two chaperones in MnP maturation and production is discussed.

Aspergillus niger↗