Retrotranscriptasas del VIH tipo 1 grupo O, activas a temperaturas elevadas

La presente invención se encuadra dentro del campo de la biotecnología. Más concretamente, se refiere a retrotranscriptasas expresadas y purificadas en bacterias y que tienen la secuencia de aminoácidos de la retrotranscriptasa de un virus de la inmunodeficiencia humana de tipo 1 (VIH-1) del grupo O modificada en las posiciones 358, 359 y 360; y de variantes de esta enzima que contienen cambios adicionales en las posiciones 355 y 357 o en la 478 o en la posición 69 (en este caso acompañada de una inserción de dos aminoácidos). Estas polimerasas presentan mayor actividad que la enzima no mutada, a temperaturas elevadas (superiores a 60ºC). Además retienen capacidad de síntesis de ADN a temperaturas superiores a 70ºC. Por otro lado, la fidelidad de copia de estas enzimas no difiere significativamente de la presentada por la retrotranscriptasa no mutada.Peer reviewedConsejo Superior de Investigaciones CientíficasA1 Solicitud de patente con informe sobre el estado de la técnic

Cross-Over between Discrete and Continuous Protein Structure Space: Insights into Automatic Classification and Networks of Protein Structures

Structural classifications of proteins assume the existence of the fold, which is an intrinsic equivalence class of protein domains. Here, we test in which conditions such an equivalence class is compatible with objective similarity measures. We base our analysis on the transitive property of the equivalence relationship, requiring that similarity of A with B and B with C implies that A and C are also similar. Divergent gene evolution leads us to expect that the transitive property should approximately hold. However, if protein domains are a combination of recurrent short polypeptide fragments, as proposed by several authors, then similarity of partial fragments may violate the transitive property, favouring the continuous view of the protein structure space. We propose a measure to quantify the violations of the transitive property when a clustering algorithm joins elements into clusters, and we find out that such violations present a well defined and detectable cross-over point, from an approximately transitive regime at high structure similarity to a regime with large transitivity violations and large differences in length at low similarity. We argue that protein structure space is discrete and hierarchic classification is justified up to this cross-over point, whereas at lower similarities the structure space is continuous and it should be represented as a network. We have tested the qualitative behaviour of this measure, varying all the choices involved in the automatic classification procedure, i.e., domain decomposition, alignment algorithm, similarity score, and clustering algorithm, and we have found out that this behaviour is quite robust. The final classification depends on the chosen algorithms. We used the values of the clustering coefficient and the transitivity violations to select the optimal choices among those that we tested. Interestingly, this criterion also favours the agreement between automatic and expert classifications. As a domain set, we have selected a consensus set of 2,890 domains decomposed very similarly in SCOP and CATH. As an alignment algorithm, we used a global version of MAMMOTH developed in our group, which is both rapid and accurate. As a similarity measure, we used the size-normalized contact overlap, and as a clustering algorithm, we used average linkage. The resulting automatic classification at the cross-over point was more consistent than expert ones with respect to the structure similarity measure, with 86% of the clusters corresponding to subsets of either SCOP or CATH superfamilies and fewer than 5% containing domains in distinct folds according to both SCOP and CATH. Almost 15% of SCOP superfamilies and 10% of CATH superfamilies were split, consistent with the notion of fold change in protein evolution. These results were qualitatively robust for all choices that we tested, although we did not try to use alignment algorithms developed by other groups. Folds defined in SCOP and CATH would be completely joined in the regime of large transitivity violations where clustering is more arbitrary. Consistently, the agreement between SCOP and CATH at fold level was lower than their agreement with the automatic classification obtained using as a clustering algorithm, respectively, average linkage (for SCOP) or single linkage (for CATH). The networks representing significant evolutionary and structural relationships between clusters beyond the cross-over point may allow us to perform evolutionary, structural, or functional analyses beyond the limits of classification schemes. These networks and the underlying clusters are available at http://ub.cbm.uam.es/research/ProtNet.phpThis work was supported by the Ramon y Cajal program of the Spanish Science Ministry of Education and Science, Project ‘Centrosoma 3DBioinformatics’ of the program Consolider-Ingenio 2010 of the Spanish Ministry of Education and Science, Project BIO2005-0576 from the Spanish Ministry of Education and Science, Project 200520M157 from the Comunidad de Madrid, and Research Foundation ‘‘Ramon Areces’’.Peer reviewe

Fluorine Abundances in the Globular Cluster M 4

We present chemical abundances for the elements carbon, sodium, and fluorine in 15 red giants of the globular cluster M 4, as well as six red giants of the globular cluster $\omega$ Centauri. The chemical abundances were calculated in LTE via spectral synthesis. The spectra analyzed are high-resolution spectra obtained in the near-infrared region around $\lambda$2.3$\mu$m with the Phoenix spectrograph on the 8.1m Gemini South Telescope, the IGRINS spectrograph on the McDonald Observatory 2.7m Telescope, and the CRIRES spectrograph on the ESO 8.2m Very Large Telescope. The results indicate a significant reduction in the fluorine abundances when compared to previous values from the literature for M 4 and $\omega$ Centauri, due to a downward revision in the excitation potentials of the HF(1-0) R9 line used in the analysis. The fluorine abundances obtained for the M 4 red giants are found to be anti-correlated with those of Na, following the typical pattern of abundance variations seen in globular clusters between distinct stellar populations. In M 4, as the Na abundance increases by $\sim$+0.4 dex, the F abundance decreases by $\sim$-0.2 dex. A comparison with abundance predictions from two sets of stellar evolution models finds that the models predict somewhat less F depletion ($\sim$-0.1 dex) for the same increase of +0.4 dex in Na

Two new alleles of the abscisic aldehyde oxidase 3 gene reveal its role in abscisic acid biosynthesis in seeds

9 páginas, 3 figuras, 3 tablas -- PAGS nros. 325-333The abscisic aldehyde oxidase 3 (AAO3) gene product of Arabidopsis catalyzes the final step in abscisic acid (ABA) biosynthesis. An aao3-1 mutant in a Landsberg erecta genetic background exhibited a wilty phenotype in rosette leaves, whereas seed dormancy was not affected (Seo et al., 2000a). Therefore, it was speculated that a different aldehyde oxidase would be the major contributor to ABA biosynthesis in seeds (Seo et al., 2000a). Through a screening based on germination under high-salt concentration, we isolated two mutants in a Columbia genetic background, initially named sre2-1 and sre2-2 (for salt resistant). Complementation tests with different ABA-deficient mutants indicated that sre2-1 and sre2-2 mutants were allelic to aao3-1, and therefore they were renamed as aao3-2 and aao3-3, respectively. Indeed, molecular characterization of the aao3-2 mutant revealed a T-DNA insertional mutation that abolished the transcription of AAO3 gene, while sequence analysis of AAO3 in aao3-3 mutant revealed a deletion of three nucleotides and several missense mutations. Physiological characterization of aao3-2 and aao3-3 mutants revealed a wilty phenotype and osmotolerance in germination assays. In contrast to aao3-1, both aao3-2 and aao3-3 mutants showed a reduced dormancy. Accordingly, ABA levels were reduced in dry seeds and rosette leaves of both aao3-2 and aao3-3. Taken together, these results indicate that AAO3 gene product plays a major role in seed ABA biosynthesisThis work was supported by the Ministerio de Ciencia y Tecnologia (grant no. BIO2002–03090) and FEDERPeer reviewe

Microarray-based identification of antigenic variants of foot-and-mouth disease virus: a bioinformatics quality assessment

BACKGROUND: The evolution of viral quasispecies can influence viral pathogenesis and the response to antiviral treatments. Mutant clouds in infected organisms represent the first stage in the genetic and antigenic diversification of RNA viruses, such as foot and mouth disease virus (FMDV), an important animal pathogen. Antigenic variants of FMDV have been classically diagnosed by immunological or RT-PCR-based methods. DNA microarrays are becoming increasingly useful for the analysis of gene expression and single nucleotide polymorphisms (SNPs). Recently, a FMDV microarray was described to detect simultaneously the seven FMDV serotypes. These results encourage the development of new oligonucleotide microarrays to probe the fine genetic and antigenic composition of FMDV for diagnosis, vaccine design, and to gain insight into the molecular epidemiology of this pathogen. RESULTS: A FMDV microarray was designed and optimized to detect SNPs at a major antigenic site of the virus. A screening of point mutants of the genomic region encoding antigenic site A of FMDV C-S8c1 was achieved. The hybridization pattern of a mutant includes specific positive and negative signals as well as crosshybridization signals, which are of different intensity depending on the thermodynamic stability of each probe-target pair. Moreover, an array bioinformatic classification method was developed to evaluate the hybridization signals. This statistical analysis shows that the procedure allows a very accurate classification per variant genome. CONCLUSION: A specific approach based on a microarray platform aimed at distinguishing point mutants within an important determinant of antigenicity and host cell tropism, namely the G-H loop of capsid protein VP1, was developed. The procedure is of general applicability as a test for specificity and discriminatory power of microarray-based diagnostic procedures using multiple oligonucleotide probes

Rubidium in the Interstellar Medium

We present observations of interstellar rubidium toward o Per, zeta Per, AE Aur, HD 147889, chi Oph, zeta Oph, and 20 Aql. Theory suggests that stable 85Rb and long-lived 87Rb are produced predominantly by high-mass stars, through a combination of the weak s- and r-processes. The 85Rb/87Rb ratio was determined from measurements of the Rb I line at 7800 angstroms and was compared to the solar system meteoritic ratio of 2.59. Within 1-sigma uncertainties all directions except HD 147889 have Rb isotope ratios consistent with the solar system value. The ratio toward HD 147889 is much lower than the meteoritic value and similar to that toward rho Oph A (Federman et al. 2004); both lines of sight probe the Rho Ophiuchus Molecular Cloud. The earlier result was attributed to a deficit of r-processed 85Rb. Our larger sample suggests instead that 87Rb is enhanced in these two lines of sight. When the total elemental abundance of Rb is compared to the K elemental abundance, the interstellar Rb/K ratio is significantly lower than the meteoritic ratio for all the sight lines in this study. Available interstellar samples for other s- and r- process elements are used to help interpret these results.Comment: 24 pages, 3 figures. Accepted for publication in Ap

Modifications in host cell structure and functions mediated by Tat intracellular expression are greatly dependent on the second exon

1 page.-- Poster presentation.Peer reviewe

Rainwater harvesting knowledge and practice for agricultural production in a changing climate: A review from Uganda’s perspective

With a changing climate in Uganda, rainfall distribution patterns have become more irregular over time and space. Excess water during rainy season is causing runoff, soil erosion, nutrient depletion and crop damage which reduce the productive capacity of land, while on the other hand, prolonged droughts during the crop growing period have become common occurrences. Additionally, pastoralists lose livestock during the dry period each year in the Cattle Corridor of Uganda due to water shortage and lack of forage. It thus remains difficult to achieve the agricultural development targets identified in the National Development Plan for Uganda, without addressing regular incidences of adverse impacts of climate change. Currently there are no well explained approaches which can contribute to adoption of technologies like rainwater management systems which are crucial in enhancing crop yields and livestock production during periods of water shortage. The overarching objective of this paper was to carry out an assessment of the status, performance, and scope for improving rainwater harvesting (RWH) for small-scale agriculture under local conditions. Accordingly, research gaps in RWH technologies were identified and documented to inform future studies. The research was carried out in the semi-arid areas of Nakasongola, Rakai, and Hoima Districts characterized by crop-livestock dependent livelihoods. Findings show that RWH Technologies can enable smallholder farmers and agro-pastoralists to become more resilient to increasing climate variability and climate change by conserving soil and water thus increasing food production and enhancing food security. Small-scale irrigation systems have enabled farmers adapt to drought challenges by enhancing crop yields and allowing farmers to target for higher market prices usually associated with the effects of drought. However, there are challenges including threats to sustainability of such established systems because of lack of community participation in systems’ monitoring and maintenances, and vandalism, and some systems require high investment costs.

Dual activation of pathways regulated by steroid receptors and peptide growth factors in primary prostate cancer revealed by Factor Analysis of microarray data

BACKGROUND: We use an approach based on Factor Analysis to analyze datasets generated for transcriptional profiling. The method groups samples into biologically relevant categories, and enables the identification of genes and pathways most significantly associated to each phenotypic group, while allowing for the participation of a given gene in more than one cluster. Genes assigned to each cluster are used for the detection of pathways predominantly activated in that cluster by finding statistically significant associated GO terms. We tested the approach with a published dataset of microarray experiments in yeast. Upon validation with the yeast dataset, we applied the technique to a prostate cancer dataset. RESULTS: Two major pathways are shown to be activated in organ-confined, non-metastatic prostate cancer: those regulated by the androgen receptor and by receptor tyrosine kinases. A number of gene markers (HER3, IQGAP2 and POR1) highlighted by the software and related to the later pathway have been validated experimentally a posteriori on independent samples. CONCLUSION: Using a new microarray analysis tool followed by a posteriori experimental validation of the results, we have confirmed several putative markers of malignancy associated with peptide growth factor signalling in prostate cancer and revealed others, most notably ERRB3 (HER3). Our study suggest that, in primary prostate cancer, HER3, together or not with HER4, rather than in receptor complexes involving HER2, could play an important role in the biology of these tumors. These results provide new evidence for the role of receptor tyrosine kinases in the establishment and progression of prostate cancer

Dual activation of pathways regulated by steroid receptors and peptide growth factors in primary prostate cancer revealed by Factor Analysis of microarray data

BACKGROUND: We use an approach based on Factor Analysis to analyze datasets generated for transcriptional profiling. The method groups samples into biologically relevant categories, and enables the identification of genes and pathways most significantly associated to each phenotypic group, while allowing for the participation of a given gene in more than one cluster. Genes assigned to each cluster are used for the detection of pathways predominantly activated in that cluster by finding statistically significant associated GO terms. We tested the approach with a published dataset of microarray experiments in yeast. Upon validation with the yeast dataset, we applied the technique to a prostate cancer dataset. RESULTS: Two major pathways are shown to be activated in organ-confined, non-metastatic prostate cancer: those regulated by the androgen receptor and by receptor tyrosine kinases. A number of gene markers (HER3, IQGAP2 and POR1) highlighted by the software and related to the later pathway have been validated experimentally a posteriori on independent samples. CONCLUSION: Using a new microarray analysis tool followed by a posteriori experimental validation of the results, we have confirmed several putative markers of malignancy associated with peptide growth factor signalling in prostate cancer and revealed others, most notably ERRB3 (HER3). Our study suggest that, in primary prostate cancer, HER3, together or not with HER4, rather than in receptor complexes involving HER2, could play an important role in the biology of these tumors. These results provide new evidence for the role of receptor tyrosine kinases in the establishment and progression of prostate cancer