Caxias do Sul, Brazil
Caxias do Sul, Brazil

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Barrero A.F.,Institute Biotecnologia | Herrador M.M.,Institute Biotecnologia | Arteaga P.,Institute Biotecnologia | Arteaga J.F.,University of Huelva | Arteaga A.F.,University of Granada
Molecules | Year: 2012

Communic acids are diterpenes with labdane skeletons found in many plant species, mainly conifers, predominating in the genus Juniperus (fam. Cupresaceae). In this review we briefly describe their distribution and different biological activities (anti- bacterial, antitumoral, hypolipidemic, relaxing smooth muscle, etc.). This paper also includes a detailed explanation of their use as chiral building blocks for the synthesis of bioactive natural products. Among other uses, communic acids have proven useful as chirons for the synthesis of quassinoids (formal), abietane antioxidants, ambrox and other perfume fixatives, podolactone herbicides, etc., featuring shorter and more efficient processes.


Geisen S.,University of Granada | Barturen G.,University of Granada | Alganza A.M.,University of Granada | Hackenberg M.,University of Granada | And 3 more authors.
Nucleic Acids Research | Year: 2014

The updated release of 'NGSmethDB' (http://bioinfo2.ugr.es/NGSmethDB) is a repository for single-base whole-genome methylome maps for the best-assembled eukaryotic genomes. Short-read data sets from NGS bisulfite-sequencing projects of cell lines, fresh and pathological tissues are first pre-processed and aligned to the corresponding reference genome, and then the cytosine methylation levels are profiled. One major improvement is the application of a unique bioinformatics protocol to all data sets, thereby assuring the comparability of all values with each other. We implemented stringent quality controls to minimize important error sources, such as sequencing errors, bisulfite failures, clonal reads or single nucleotide variants (SNVs). This leads to reliable and high-quality methylomes, all obtained under uniform settings. Another significant improvement is the detection in parallel of SNVs, which might be crucial for many downstream analyses (e.g. SNVs and differential-methylation relationships). A next-generation methylation browser allows fast and smooth scrolling and zooming, thus speeding data download/upload, at the same time requiring fewer server resources. Several data mining tools allow the comparison/retrieval of methylation levels in different tissues or genome regions. NGSmethDB methylomes are also available as native tracks through a UCSC hub, which allows comparison with a wide range of third-party annotations, in particular phenotype or disease annotations. © 2013 The Author(s). Published by Oxford University Press.


Bernal D.,University of Valencia | Trelis M.,University of Valencia | Montaner S.,University of Valencia | Cantalapiedra F.,University of Valencia | And 4 more authors.
Journal of Proteomics | Year: 2014

With the aim of characterizing the molecules involved in the interaction of Dicrocoelium dendriticum adults and the host, we have performed proteomic analyses of the external surface of the parasite using the currently available datasets including the transcriptome of the related species Echinostoma caproni. We have identified 182 parasite proteins on the outermost surface of D. dendriticum. The presence of exosome-like vesicles in the ESP of D. dendriticum and their components has also been characterized. Using proteomic approaches, we have characterized 84 proteins in these vesicles. Interestingly, we have detected miRNA in D. dendriticum exosomes, thus representing the first report of miRNA in helminth exosomes. Biological significance: In order to identify potential targets for intervention against parasitic helminths, we have analyzed the surface of the parasitic helminth Dicrocoelium dendriticum. Along with the proteomic analyses of the outermost layer of the parasite, our work describes the molecular characterization of the exosomes of D. dendriticum. Our proteomic data confirm the improvement of protein identification from "non-model organisms" like helminths, when using different search engines against a combination of available databases. In addition, this work represents the first report of miRNAs in parasitic helminth exosomes. These vesicles can pack specific proteins and RNAs providing stability and resistance to RNAse digestion in body fluids, and provide a way to regulate host-parasite interplay. The present data should provide a solid foundation for the development of novel methods to control this non-model organism and related parasites. This article is part of a Special Issue entitled: Proteomics of non-model organisms. © 2014 Elsevier B.V.


Centeno D.C.,Max Planck Institute of Molecular Plant Physiology | Osorio S.,Max Planck Institute of Molecular Plant Physiology | Nunes-Nesi A.,Max Planck Institute of Molecular Plant Physiology | Bertolo A.L.F.,Cornell University | And 11 more authors.
Plant Cell | Year: 2011

Despite the fact that the organic acid content of a fruit is regarded as one of its most commercially important quality traits when assessed by the consumer, relatively little is known concerning the physiological importance of organic acid metabolism for the fruit itself. Here, we evaluate the effect of modifying malate metabolism in a fruit-specific manner, by reduction of the activities of either mitochondrial malate dehydrogenase or fumarase, via targeted antisense approaches in tomato (Solanum lycopersicum). While these genetic perturbations had relatively little effect on the total fruit yield, they had dramatic consequences for fruit metabolism, as well as unanticipated changes in postharvest shelf life and susceptibility to bacterial infection. Detailed characterization suggested that the rate of ripening was essentially unaltered but that lines containing higher malate were characterized by lower levels of transitory starch and a lower soluble sugars content at harvest, whereas those with lower malate contained higher levels of these carbohydrates. Analysis of the activation state of ADP-glucose pyrophosphorylase revealed that it correlated with the accumulation of transitory starch. Taken together with the altered activation state of the plastidial malate dehydrogenase and the modified pigment biosynthesis of the transgenic lines, these results suggest that the phenotypes are due to an altered cellular redox status. The combined data reveal the importance of malate metabolism in tomato fruit metabolism and development and confirm the importance of transitory starch in the determination of agronomic yield in this species. © American Society of Plant Biologists.


Nahirnak V.,Institute Biotecnologia
Plant signaling & behavior | Year: 2012

Snakin/GASA proteins are widely distributed among plant species. They are expressed in different plant organs with high tissue and temporal specificity, and their subcellular localization varies among the different members. Interestingly, all of them maintain 12 cysteines of the C-terminus in highly conserved positions of the aminoacid sequences that are essential for their biochemical activity and probably responsible for their protein structure. Despite their common features, their functions are not completely elucidated and little is known about their mode of action. This review focuses on the current knowledge about this intriguing family of peptides and advances comprising gene regulation analyses, expression pattern studies and phenotypic characterization of mutants and transgenic plants. Furthermore, we discuss the roles of Snakin/GASA proteins in several aspects of plant development, plant responses to biotic or abiotic stress and their participation in hormone crosstalk and redox homeostasis.


Nahirnak V.,Institute Biotecnologia | Almasia N.I.,Institute Biotecnologia | Hopp H.E.,Institute Biotecnologia | Vazquez-Rovere C.,Institute Biotecnologia
Plant Signaling and Behavior | Year: 2012

Snakin/GASA proteins are widely distributed among plant species. They are expressed in different plant organs with high tissue and temporal specificity, and their subcellular localization varies among the different members. interestingly, all of them maintain 12 cysteines of the C-terminus in highly conserved positions of the aminoacid sequences that are essential for their biochemical activity and probably responsible for their protein structure. Despite their common features, their functions are not completely elucidated and little is known about their mode of action. This review focuses on the current knowledge about this intriguing family of peptides and advances comprising gene regulation analyses, expression pattern studies and phenotypic characterization of mutants and transgenic plants. Furthermore, we discuss the roles of Snakin/GASA proteins in several aspects of plant development, plant responses to biotic or abiotic stress and their participation in hormone crosstalk and redox homeostasis. © 2012 Landes Bioscience.


Guerrero-Olazaran M.,Institute Biotecnologia | Rodriguez-Blanco L.,Institute Biotecnologia | Carreon-Trevino J.G.,Institute Biotecnologia | Gallegos-Lopez J.A.,Institute Biotecnologia | Viader-Salvado J.M.,Institute Biotecnologia
Applied and Environmental Microbiology | Year: 2010

The cloning and expression of a native gene encoding a Bacillus subtilis phytase using Pichia pastoris as the host is described. In addition, the influence of N-glycosylation on the biochemical properties of the B. subtilis phytase, the influence of pH on the thermostability of the recombinant and native B. subtilis phytases, and the resistance of both phytases to shrimp digestive enzymes and porcine trypsin are also described. After 48 h of methanol induction in shake flasks, a selected recombinant strain produced and secreted 0.82 U/ml (71 mg/liter) recombinant phytase. This phytase was N-glycosylated, had a molecular mass of 39 kDa after N-deglycosylation, exhibited activity within a pH range of 2.5 to 9 and at temperatures of 25 to 70°C, had high residual activity (85% ± 2%) after 10 min of heat treatment at 80°C and pH 5.5 in the presence of 5 mM CaCl2, and was resistant to shrimp digestive enzymes and porcine trypsin. Although the recombinant Bacillus phytase had pH and temperature activity profiles that were similar to those of the corresponding nonglycosylated native phytase, the thermal stabilities of the recombinant and native phytases were different, although both were calcium concentration and pH dependent. Copyright © 2010, American Society for Microbiology. All Rights Reserved.


Megia-Fernandez A.,Institute Biotecnologia | Ortega-Munoz M.,Institute Biotecnologia | Lopez-Jaramillo J.,Institute Biotecnologia | Hernandez-Mateo F.,Institute Biotecnologia | Santoyo-Gonzalez F.,Institute Biotecnologia
Advanced Synthesis and Catalysis | Year: 2010

Novel supported chelating adsorbents bearing diverse multidentate nitrogenated ligands with strong copper(I) affinities are easily prepared in non-magnetic and magnetic variants using silica and silica-coated magnetite nanoparticles as suitable supports and the aza-Michael-type addition of vinyl sulfones as the ligation tool. These adsorbents are versatile materials with applications in the copper-catalyzed alkyne-azide cycloaddition (CuAAC) click chemistry where their complexation abilities enable them to act either as heterogeneous click catalysts when used in their complexed form or as copper(I) scavengers when used in their uncomplexed form. In the first instance, they proved to be robust and efficient heterogeneous catalysts to promote click reactions using extremely low doses and showing negligible copper leaching, particularly in the case of the silica-based non-magnetic adsorbents, allowing a simple operational protocol for their rapid and easy removal by filtration or magnetic decantation and showing good recyclability properties. In their uncomplexed form, the non-magnetic chelating adsorbents are very efficient copper scavengers that are able to remove any traces of metal contamination and that can be applied in tandem with any heterogeneous supported copper(I) catalysts or as stand-alone copper removing system in any click protocol allowing the isolation of metal-free clicked compounds. Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Robles-Reglero V.,University of León | Robles-Reglero V.,Institute Biotecnologia | Santamarta I.,University of León | Santamarta I.,Institute Biotecnologia | And 6 more authors.
Journal of Biotechnology | Year: 2013

Expression of the holomycin biosynthesis genes (hlm) has been studied in the wild type strain Streptomyces clavuligerus ATCC 27064 and holomycin overproducer mutants. RT-PCR transcription analysis of S. clavuligerus oppA2::aph showed a higher transcription of the hlmA, B, C, D, E, F, G, H, I and hlmL genes, a slightly lower expression for hlmK and no significant differences for the transcription of the two putative regulatory genes, hlmM and hlmJ, in relation to the wild type strain. Accordingly, protein spots corresponding to HlmD, HlmF and HlmG, which were barely detectable in the wild type strain, were present in high amounts in the holomycin overproducer S. clavuligerus oppA2::aph proteome. Transcription start point analysis of the hlm genes revealed that the annotated sequences in the databases for several hlm genes were incorrect. The hlm cluster was introduced into Streptomyces coelicolor M1154 and holomycin production by S. coelicolor M1154 [pVR- hol1] was validated by bioassays and confirmed by HPLC analysis and mass spectrometry. Heterologous holomycin production by the S. coelicolor transformant is 500-fold lower than in S. clavuligerus oppA2::aph. The transformant S. coelicolor M1154 [pVR-hol1] shows holomycin sensitivity to 100 μg/ml, similar to that of the parental S. coelicolor M1154 strain, suggesting that heterologous expression in S. coelicolor might be toxic due to the lack of an holomycin resistance gene in this host strain. © 2012 Elsevier B.V.


Lopez-Garcia M.T.,University of León | Lopez-Garcia M.T.,Institute Biotecnologia | Santamarta I.,University of León | Santamarta I.,Institute Biotecnologia | And 2 more authors.
Microbiology | Year: 2010

The TTA codon-containing adpA gene of Streptomyces clavuligerus, located upstream of ornA, is in a DNA region syntenous with the homologous region of other Streptomyces genomes. Deletion of adpA results in a medium-dependent sparse aerial mycelium formation and lack of sporulation. Clavulanic acid formation in this mutant decreases to about 10% of the wild-type level depending on the medium, whereas its production is strongly stimulated by increasing the adpA copy number. Quantitative transcriptional analysis indicates that expression of the clavulanic acid regulatory genes ccaR and claR decreases seven- and fourfold, respectively, in the ΔadpA mutant, resulting in a large decrease in expression of genes encoding biosynthesis enzymes for the early steps of clavulanic acid formation and a smaller decrease in the expression of genes for the late steps of the pathway. An ARE box, 5′-TCTCATGGAGACATAGCGGGGCATGC-3′, is present upstream of adpA and efficiently binds S. clavuligerus Brp protein, as shown by electrophoretic mobility shift assay (EMSA) analysis. The transcription level of adpA is higher in the absence of Brp, as shown in S. clavuligerus Δbrp, suggesting a connection between adpA expression and the c-butyrolactone system in S. clavuligerus. © 2010 SGM.

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