20 research outputs found
Presentación del monográfico Trabajo Social y Economía
Presentación del monográfico Trabajo Social y Economí
Presentación del monográfico Trabajo Social y Economía
Presentación del monográfico Trabajo Social y Economí
Study of the role played by NfsA, NfsB nitroreductase and NemA flavin reductase from Escherichia coli in the conversion of ethyl 2-(2′-nitrophenoxy)acetate to 4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one (D-DIBOA), a benzohydroxamic acid with interesting biological properties
Benzohydroxamic acids, such as 4-hydroxy-(2H)- 1,4-benzoxazin-3(4H)-one (D-DIBOA), exhibit interesting herbicidal, fungicidal and bactericidal properties. Recently, the chemical synthesis of D-DIBOA has been simplified to only two steps. In a previous paper, we demonstrated that the second step could be replaced by a biotransformation using Escherichia coli to reduce the nitro group of the precursor, ethyl 2-(2′-nitrophenoxy)acetate and obtain D-DIBOA. The
NfsA and NfsB nitroreductases and the NemA xenobiotic reductase of E. coli have the capacity to reduce one or two
nitro groups from a wide variety of nitroaromatic compounds, which are similar to the precursor. By this reason, we hypothesised that these three enzymes could be involved in this biotransformation. We have analysed the biotransformation yield (BY) of mutant strains in which one, two or three of these genes were knocked out, showing that only in the double nfsA/nfsB and in the triple nfsA/nfsB/nemA mutants, the BY was 0%. These results suggested that NfsA and NfsB are responsible for the biotransformation in the tested conditions. To confirm this, the nfsA and nfsB open reading frames were cloned into the pBAD expression vector and transformed into the nfsA and nfsB single mutants, respectively. In both cases, the biotransformation capacity of the strains was recovered (6.09±0.06% as in the wild-type strain) and incremented considerably when NfsA and NfsB were overexpressed (40.33%±9.42% and 59.68%±2.0% respectively)
Identification of Enzymatic Bottlenecks for the Aerobic Production of Malate from Glycerol by the Systematic Gene Overexpression of Anaplerotic Enzymes in Escherichia coli
The biotechnological production of dicarboxylic acids (C4) from renewable carbon sources represents an attractive approach for the provision of these valuable compounds by green chemistry means. Glycerol has become a waste product of the biodiesel industry that serves as a highly reduced carbon source for some microorganisms. Escherichia coli is capable of consuming glycerol to produce succinate under anaerobic fermentation, but with the deletion of some tricarboxylic acid (TCA) cycle genes, it is also able to produce succinate and malate in aerobiosis. In this study, we investigate possible rate-limiting enzymes by overexpressing the C-feeding anaplerotic enzymes Ppc, MaeA, MaeB, and Pck in a mutant that lacks the succinate dehydrogenase (Sdh) enzyme. The overexpression of the TCA enzyme Mdh and the activation of the glyoxylate shunt was also examined. Using this unbiased approach, we found that phosphoenol pyruvate carboxylase (Ppc) overexpression enhances an oxidative pathway that leads to increasing succinate, while phosphoenol pyruvate carboxykinase (Pck) favors a more efficient reductive branch that produces mainly malate, at 57.5% of the theoretical maximum molar yield. The optimization of the culture medium revealed the importance of bicarbonate and pH in the production of malate. An additional mutation of the ppc gene highlights its central role in growth and C4 production
Optimization of the Biocatalysis for D-DIBOA Synthesis Using a Quick and Sensitive New Spectrophotometric Quantification Method
D-DIBOA (4-hydroxy-(2H)-1,4-benzoxazin-3-(4H)-one) is an allelopathic-derived compound with interesting herbicidal, fungicidal, and insecticide properties whose production has been successfully achieved by biocatalysis using a genetically engineered Escherichia coli strain. However, improvement and scaling-up of this process are hampered by the current methodology for D-DIBOA quantification, which is based on high-performance liquid chromatographic (HPLC), a time-consuming technique that requires expensive equipment and the use of environmentally unsafe solvents. In this work, we established and validated a rapid, simple, and sensitive spectrophotometric method for the quantification of the D-DIBOA produced by whole-cell biocatalysis, with limits of detection and quantification of 0.0165 and 0.0501 mu mol center dot mL(-1) respectively. This analysis takes place in only a few seconds and can be carried out using 100 mu L of the sample in a microtiter plate reader. We performed several whole-cell biocatalysis strategies to optimize the process by monitoring D-DIBOA production every hour to keep control of both precursor and D-DIBOA concentrations in the bioreactor. These experiments allowed increasing the D-DIBOA production from the previously reported 5.01 mM up to 7.17 mM (43% increase). This methodology will facilitate processes such as the optimization of the biocatalyst, the scaling up, and the downstream purification
A systematic analysis of TCA Escherichia coli mutants reveals suitable genetic backgrounds for enhanced hydrogen and ethanol production using glycerol as main carbon source
Biodiesel has emerged as an environmentally friendly alternative to fossil fuels; however, the low price of glycerol feed-stocks generated from the biodiesel industry has become a burden to this industry. A feasible alternative is the microbial biotransformation of waste glycerol to hydrogen and ethanol. Escherichia coli, a microorganism commonly used for metabolic engineering, is able to biotransform glycerol into these products. Nevertheless, the wild type strain yields can be improved by rewiring the carbon flux to the desired products by genetic engineering. Due to the importance of the central carbon metabolism in hydrogen and ethanol synthesis, E. coli single null mutant strains for enzymes of the TCA cycle and other related reactions were studied in this work. These strains were grown anaerobically in a glycerol-based medium and the concentrations of ethanol, glycerol, succinate and hydrogen were analysed by HPLC and GC. It was found that the reductive branch is the more relevant pathway for the aim of this work, with malate playing a central role. It was also found that the putative C4-transporter dcuD mutant improved the target product yields. These results will contribute to reveal novel metabolic engineering strategies for improving
hydrogen and ethanol production by E. coli
Automatable downstream purification of the benzohydroxamic acid D-DIBOA from a biocatalytic synthesis
Herbicides play a vital role in agriculture, contributing to increased crop productivity by minimizing weed growth, but their low degradability presents a threat to the environment and human health. Allelochemicals, such as DIBOA (2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4 H)-one), are secondary metabolites released by certain plants that affect the survival or growth of other organisms. Although these metabolites have an attractive po-tential for use as herbicides, their low natural production is a critical hurdle. Previously, the synthesis of the biologically active analog D-DIBOA (4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one) was achieved, using an engi-neered E. coli strain as a whole-cell biocatalyst, capable of transforming a precursor compound into D-DIBOA and exporting it into the culture medium, although it cannot be directly applied to crops. Here a chromatographic method to purify D-DIBOA from this cell culture medium without producing organic solvent wastes is described. The purification of D-DIBOA from a filtered culture medium to the pure compound could also be automated. Biological tests with the purified compound on weed models showed that it has virtually the same activity than the chemically synthesized D-DIBOA
Co-occurrence of colistin-resistance genes mcr-1 and mcr-3 among multidrug-resistant Escherichia coli isolated from cattle, Spain, September 2015
Los genes de resistencia a colistina mcr-3 y mcr-1 se detectaron en un aislado de Escherichia coli de heces de ganado en un matadero español en 2015. Las secuencias de ambos genes se hibridaron a la misma banda de plásmidos de aproximadamente 250 kb, aunque la resistencia a la colistina no fue movilizable . El aislado producía betalactamasas de espectro extendido y pertenecía al serotipo O9: H10 y al tipo de secuencia ST533. Aquí informamos un gen mcr-3 detectado en Europa después de informes anteriores de Asia y los Estados Unidos.Colistin resistance genes mcr-3 and mcr-1 have been detected in an Escherichia coli isolate from cattle faeces in a Spanish slaughterhouse in 2015. The sequences of both genes hybridised to same plasmid band of ca 250 kb, although colistin resistance was non-mobilisable. The isolate was producing extended-spectrum beta-lactamases and belonged to serotype O9:H10 and sequence type ST533. Here we report an mcr-3 gene detected in Europe following earlier reports from Asia and the United States.• Ministerio de Economía, Industria y Competitividad. Proyecto AGL2016-
74882-C3
• Ministerio de Agricultura y Pesca (España) y Comunidad Autónoma de Comunidad Autónoma de Madrid. Ayuda S2013 / ABI-2747
• Junta de Extremadura y Fondo Europeo de Desarrollo Regional. Ayuda GR15075 e IB16073
• Fundación para la Ciencia y la Tecnología (Portugal). Ayudas UID / MAR / 04292/2013
• Fundación Tatiana de Guzmán El Bueno (España). Beca doctoral para María del Rocío Iglesias Parro
• Instituto Nacional de Agricultura y Alimentación. Investigación y Tecnología (INIA). Beca doctoral para María del Rocío Iglesias Parro
• Ministerio de Economía, Industria y Competitividad. Beca FPI2014-020, para Narciso Martín QuijadapeerReviewe
Brentuximab vedotin in the treatment of cutaneous T-cell lymphomas: Data from the Spanish Primary Cutaneous Lymphoma Registry
[Background] Brentuximab vedotin (BV) has been approved for CD30-expressing cutaneous T-cell lymphoma (CTCL) after at least one previous systemic treatment. However, real clinical practice is still limited.[Objectives] To evaluate the response and tolerance of BV in a cohort of patients with CTCL.[Methods] We analysed CTCL patients treated with BV from the Spanish Primary Cutaneous Lymphoma Registry (RELCP).[Results] Sixty-seven patients were included. There were 26 females and the mean age at diagnosis was 59 years. Forty-eight were mycosis fungoides (MF), 7 Sézary syndrome (SS) and 12 CD30+ lymphoproliferative disorders (CD30 LPD). Mean follow-up was 18 months. Thirty patients (45%) showed at least 10% of CD30+ cells among the total lymphocytic infiltrate. The median number of BV infusions received was 7. The overall response rate (ORR) was 67% (63% in MF, 71% in SS and 84% in CD30 LPD). Ten of 14 patients with folliculotropic MF (FMF) achieved complete or partial response (ORR 71%). The median time to response was 2.8 months. During follow-up, 36 cases (54%) experienced cutaneous relapse or progression. The median progression free survival (PFS) was 10.3 months. The most frequent adverse event was peripheral neuropathy (PN) (57%), in most patients (85%), grades 1 or 2.[Conclusions] These results confirm the efficacy and safety of BV in patients with advanced-stage MF, and CD30 LPD. In addition, patients with FMF and SS also showed a favourable response. Our data suggest that BV retreatment is effective in a proportion of cases.The Spanish Primary Cutaneous Lymphoma Registry (RELCP) is promoted by the Fundación Piel Sana Academia Española de Dermatología y Venereología, which received an unrestricted grant support from Kyowa Kirin.Peer reviewe
RICORS2040 : The need for collaborative research in chronic kidney disease
Chronic kidney disease (CKD) is a silent and poorly known killer. The current concept of CKD is relatively young and uptake by the public, physicians and health authorities is not widespread. Physicians still confuse CKD with chronic kidney insufficiency or failure. For the wider public and health authorities, CKD evokes kidney replacement therapy (KRT). In Spain, the prevalence of KRT is 0.13%. Thus health authorities may consider CKD a non-issue: very few persons eventually need KRT and, for those in whom kidneys fail, the problem is 'solved' by dialysis or kidney transplantation. However, KRT is the tip of the iceberg in the burden of CKD. The main burden of CKD is accelerated ageing and premature death. The cut-off points for kidney function and kidney damage indexes that define CKD also mark an increased risk for all-cause premature death. CKD is the most prevalent risk factor for lethal coronavirus disease 2019 (COVID-19) and the factor that most increases the risk of death in COVID-19, after old age. Men and women undergoing KRT still have an annual mortality that is 10- to 100-fold higher than similar-age peers, and life expectancy is shortened by ~40 years for young persons on dialysis and by 15 years for young persons with a functioning kidney graft. CKD is expected to become the fifth greatest global cause of death by 2040 and the second greatest cause of death in Spain before the end of the century, a time when one in four Spaniards will have CKD. However, by 2022, CKD will become the only top-15 global predicted cause of death that is not supported by a dedicated well-funded Centres for Biomedical Research (CIBER) network structure in Spain. Realizing the underestimation of the CKD burden of disease by health authorities, the Decade of the Kidney initiative for 2020-2030 was launched by the American Association of Kidney Patients and the European Kidney Health Alliance. Leading Spanish kidney researchers grouped in the kidney collaborative research network Red de Investigación Renal have now applied for the Redes de Investigación Cooperativa Orientadas a Resultados en Salud (RICORS) call for collaborative research in Spain with the support of the Spanish Society of Nephrology, Federación Nacional de Asociaciones para la Lucha Contra las Enfermedades del Riñón and ONT: RICORS2040 aims to prevent the dire predictions for the global 2040 burden of CKD from becoming true