RESUMO
The use of fossil fuels has been strongly related to critical problems currently affecting society, such as: global warming, global greenhouse effects and pollution. These problems have affected the homeostasis of living organisms worldwide at an alarming rate. Due to this, it is imperative to look for alternatives to the use of fossil fuels and one of the relevant substitutes are biofuels. There are different types of biofuels (categories and generations) that have been previously explored, but recently, the use of microalgae has been strongly considered for the production of biofuels since they present a series of advantages over other biofuel production sources: (a) they don't need arable land to grow and therefore do not compete with food crops (like biofuels produced from corn, sugar cane and other plants) and; (b) they exhibit rapid biomass production containing high oil contents, at least 15 to 20 times higher than land based oleaginous crops. Hence, these unicellular photosynthetic microorganisms have received great attention from researches to use them in the large-scale production of biofuels. However, one disadvantage of using microalgae is the high economic cost due to the low-yields of lipid content in the microalgae biomass. Thus, development of different methods to enhance microalgae biomass, as well as lipid content in the microalgae cells, would lead to the development of a sustainable low-cost process to produce biofuels. Within the last 10 years, many studies have reported different methods and strategies to induce lipid production to obtain higher lipid accumulation in the biomass of microalgae cells; however, there is not a comprehensive review in the literature that highlights, compares and discusses these strategies. Here, we review these strategies which include modulating light intensity in cultures, controlling and varying CO2 levels and temperature, inducing nutrient starvation in the culture, the implementation of stress by incorporating heavy metal or inducing a high salinity condition, and the use of metabolic and genetic engineering techniques coupled with nanotechnology.
Assuntos
Biocombustíveis , Lipídeos/biossíntese , Engenharia Metabólica/métodos , Microalgas , Fermentação , Microalgas/genética , Microalgas/crescimento & desenvolvimento , Microalgas/metabolismo , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/metabolismoRESUMO
Leishmaniasis is a parasitic infection caused by several species of the genus Leishmania that is considered as a neglected disease. Drug development process requires a robust and updated high-throughput technology to the evaluation of candidate compounds that imply the manipulation of the pathogenic species of the parasite in the laboratory. Therefore, it is restricted to trained personal and level II biosafety environments. However, it has been established the utility of Leishmania tarentolae as a model for in vitro screening of antileishmanial agents without the necessity of level II biosafety setups. In parallel the transfection of Leishmania parasites with reporter genes as the eGFP using non-commercial integration vectors like the pIRmcs3(-) has proved to be a powerful tool for the implementation of semi automatized high-throughput platforms for the evaluation of antileishmanial compounds. Here we report the generation of a new L. tarentolae strain overexpressing the eGFP gene harboured by the non-commercial vector pIR3(-). We also demonstrate its utility for the semi-automatized screening of antileshmanial compounds in intracellular forms of the L. tarentolae parasite.
Assuntos
Antiprotozoários/farmacologia , Leishmania/genética , Leishmaniose/parasitologia , Doenças Negligenciadas/parasitologia , Organismos Geneticamente Modificados/genética , Antiprotozoários/uso terapêutico , Biologia Computacional , Descoberta de Drogas , Citometria de Fluxo , Expressão Gênica , Genes Reporter , Vetores Genéticos , Proteínas de Fluorescência Verde/genética , Ensaios de Triagem em Larga Escala , Humanos , Concentração Inibidora 50 , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Doenças Negligenciadas/tratamento farmacológico , RNA Ribossômico 18S/genética , Segurança , Células U937RESUMO
To facilitate the biodegradation of diesel oil, an oil biodegradation bacterial consortium was constructed. The alkane hydroxylase (alkB) gene of Pseudomonas putida GPo1 was constructed in a pCom8 expression vector, and the pCom8-GPo1 alkB plasmid was transformed into Escherichia coli DH5α. The AlkB protein was expressed by diesel oil induction and detected through SDS-polyacrylamide gel electrophoresis. The culture of the recombinant (pCom8-GPo1 alkB/E. coli DH5α) with the oil biodegradation bacterial consortium increased the degradation ratio of diesel oil at 24 h from 31% to 50%, and the facilitation rates were increased as the proportion of pCom8-GPo1 alkB/E. coli DH5α to the consortium increased. The results suggested that the expression of the GPo1 gene in E. coli DH5α could enhance the function of diesel oil degradation by the bacterial consortium.
Assuntos
Acinetobacter/metabolismo , Biodegradação Ambiental , Citocromo P-450 CYP4A/genética , Escherichia coli/metabolismo , Consórcios Microbianos/genética , Organismos Geneticamente Modificados/metabolismo , Pseudomonas putida/enzimologia , Acinetobacter/genética , Escherichia coli/enzimologia , Escherichia coli/genética , Óleos Combustíveis , Gasolina , Engenharia Genética , Organismos Geneticamente Modificados/genética , Oxirredução , Plasmídeos/genética , Pseudomonas putida/genética , Pseudomonas putida/metabolismoRESUMO
To facilitate the biodegradation of diesel oil, an oil biodegradation bacterial consortium was constructed. The alkane hydroxylase (
Assuntos
Acinetobacter/metabolismo , Biodegradação Ambiental , /genética , Escherichia coli/metabolismo , Consórcios Microbianos/genética , Organismos Geneticamente Modificados/metabolismo , Pseudomonas putida/enzimologia , Acinetobacter/genética , Escherichia coli/enzimologia , Escherichia coli/genética , Óleos Combustíveis , Gasolina , Engenharia Genética , Oxirredução , Organismos Geneticamente Modificados/genética , Plasmídeos/genética , Pseudomonas putida/genética , Pseudomonas putida/metabolismoRESUMO
Cytochrome c oxidase (CcO) biogenesis requires several accessory proteins implicated, among other processes, in copper and haem a insertion. In yeast, the farnesyltransferase Cox10p that catalyses the conversion of haem b to haem o is the limiting factor in haem a biosynthesis and is essential for haem a insertion in CcO. In this work, we characterized AtCOX10, a putative Cox10p homologue from Arabidopsis thaliana. AtCOX10 was localized in mitochondria and was able to restore growth of a yeast Δcox10 null mutant on non-fermentable carbon sources, suggesting that it also participates in haem o synthesis. Plants with T-DNA insertions in the coding region of both copies of AtCOX10 could not be recovered, and heterozygous mutant plants showed seeds with embryos arrested at early developmental stages that lacked CcO activity. Heterozygous mutant plants exhibited lower levels of CcO activity and cyanide-sensitive respiration but normal levels of total respiration at the expense of an increase in alternative respiration. AtCOX10 seems to be implicated in the onset and progression of senescence, since heterozygous mutant plants showed a faster decrease in chlorophyll content and photosynthetic performance than wild-type plants after natural and dark-induced senescence. Furthermore, complementation of mutants by expressing AtCOX10 under its own promoter allowed us to obtain plants with T-DNA insertions in both AtCOX10 copies, which showed phenotypic characteristics comparable to those of wild type. Our results highlight the relevance of haem o synthesis in plants and suggest that this process is a limiting factor that influences CcO activity levels, mitochondrial respiration, and plant senescence.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Farnesiltranstransferase/genética , Heme/metabolismo , Proteínas Mitocondriais/genética , Arabidopsis/embriologia , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Respiração Celular , Farnesiltranstransferase/metabolismo , Proteínas Mitocondriais/metabolismo , Organismos Geneticamente Modificados/genética , Saccharomyces cerevisiae/genéticaRESUMO
Genetically attenuated microorganisms, pathogens, and some commensal bacteria can be engineered to deliver recombinant heterologous antigens to stimulate the host immune system, while still offering good levels of safety. A key feature of these live vectors is their capacity to stimulate mucosal as well as humoral and/or cellular systemic immunity. This enables the use of different forms of vaccination to prevent pathogen colonization of mucosal tissues, the front door for many infectious agents. Furthermore, delivery of DNA vaccines and immune system stimulatory molecules, such as cytokines, can be achieved using these special carriers, whose adjuvant properties and, sometimes, invasive capacities enhance the immune response. More recently, the unique features and versatility of these vectors have also been exploited to develop anti-cancer vaccines, where tumor-associated antigens, cytokines, and DNA or RNA molecules are delivered. Different strategies and genetic tools are constantly being developed, increasing the antigenic potential of agents delivered by these systems, opening fresh perspectives for the deployment of vehicles for new purposes. Here we summarize the main characteristics of the different types of live bacterial vectors and discuss new applications of these delivery systems in the field of vaccinology.
Assuntos
Animais , Humanos , Vacinas Bacterianas/imunologia , Portadores de Fármacos , Infecções Bacterianas/prevenção & controle , Vacinas Bacterianas/genética , Neoplasias/terapia , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologiaRESUMO
Genetically attenuated microorganisms, pathogens, and some commensal bacteria can be engineered to deliver recombinant heterologous antigens to stimulate the host immune system, while still offering good levels of safety. A key feature of these live vectors is their capacity to stimulate mucosal as well as humoral and/or cellular systemic immunity. This enables the use of different forms of vaccination to prevent pathogen colonization of mucosal tissues, the front door for many infectious agents. Furthermore, delivery of DNA vaccines and immune system stimulatory molecules, such as cytokines, can be achieved using these special carriers, whose adjuvant properties and, sometimes, invasive capacities enhance the immune response. More recently, the unique features and versatility of these vectors have also been exploited to develop anti-cancer vaccines, where tumor-associated antigens, cytokines, and DNA or RNA molecules are delivered. Different strategies and genetic tools are constantly being developed, increasing the antigenic potential of agents delivered by these systems, opening fresh perspectives for the deployment of vehicles for new purposes. Here we summarize the main characteristics of the different types of live bacterial vectors and discuss new applications of these delivery systems in the field of vaccinology.
Assuntos
Humanos , Animais , Vacinas Bacterianas/imunologia , Portadores de Fármacos , Infecções Bacterianas/prevenção & controle , Vacinas Bacterianas/genética , Neoplasias/terapia , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologiaRESUMO
Natural and modified nucleoside-5'-monophosphates and their precursors are valuable compounds widely used in biochemical studies. Bacterial nonspecific acid phosphatases (NSAPs) are a group of enzymes involved in the hydrolysis of phosphoester bonds, and some of them exhibit phosphotransferase activity. NSAP containing Enterobacter aerogenes and Raoultella planticola whole cells were evaluated in the phosphorylation of a wide range of nucleosides and nucleoside precursors using pyrophosphate as phosphate donor. To increase the productivity of the process, we developed two genetically modified strains of Escherichia coli which overexpressed NSAPs of E. aerogenes and R. planticola. These new recombinant microorganisms (E. coli BL21 pET22b-phoEa and E. coli BL21 pET22b-phoRp) showed higher activity than the corresponding wild-type strains. Reductions in the reaction times from 21 h to 60 min, from 4 h to 15 min, and from 24 h to 40 min in cases of dihydroxyacetone, inosine, and fludarabine, respectively, were obtained.
Assuntos
Fosfatase Ácida/metabolismo , Metabolismo dos Carboidratos , Enterobacteriaceae/enzimologia , Nucleosídeos/metabolismo , Fosfotransferases/metabolismo , Enterobacteriaceae/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/metabolismo , Fatores de TempoRESUMO
Genetically attenuated microorganisms, pathogens, and some commensal bacteria can be engineered to deliver recombinant heterologous antigens to stimulate the host immune system, while still offering good levels of safety. A key feature of these live vectors is their capacity to stimulate mucosal as well as humoral and/or cellular systemic immunity. This enables the use of different forms of vaccination to prevent pathogen colonization of mucosal tissues, the front door for many infectious agents. Furthermore, delivery of DNA vaccines and immune system stimulatory molecules, such as cytokines, can be achieved using these special carriers, whose adjuvant properties and, sometimes, invasive capacities enhance the immune response. More recently, the unique features and versatility of these vectors have also been exploited to develop anti-cancer vaccines, where tumor-associated antigens, cytokines, and DNA or RNA molecules are delivered. Different strategies and genetic tools are constantly being developed, increasing the antigenic potential of agents delivered by these systems, opening fresh perspectives for the deployment of vehicles for new purposes. Here we summarize the main characteristics of the different types of live bacterial vectors and discuss new applications of these delivery systems in the field of vaccinology.
Assuntos
Vacinas Bacterianas/imunologia , Portadores de Fármacos , Animais , Infecções Bacterianas/prevenção & controle , Vacinas Bacterianas/genética , Humanos , Neoplasias/terapia , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologiaRESUMO
Trypanosoma cruzi is the etiological agent of Chagas disease, an illness that affects about 10 million people, mostly in South America, for which there is no effective treatment or vaccine. In this context, transgenic parasites expressing reporter genes are interesting tools for investigating parasite biology and host-parasite interactions, with a view to developing new strategies for disease prevention and treatment. We describe here the construction of a stably transfected fluorescent T. cruzi clone in which the GFP gene is integrated into the chromosome carrying the ribosomal cistron in T. cruzi Dm28c. This fluorescent T. cruzi produces detectable amounts of GFP only at replicative stages (epimastigote and amastigote), consistent with the larger amounts of GFP mRNA detected in these forms than in the non replicative trypomastigote stages. The fluorescence signal was also strongly correlated with the total number of parasites in T. cruzi cultures, providing a simple and rapid means of determining the growth inhibitory dose of anti-T.cruzi drugs in epimastigotes, by fluorometric microplate screening, and in amastigotes, by the flow cytometric quantification of T. cruzi-infected Vero cells. This fluorescent T. cruzi clone is, thus, an interesting tool for unbiased detection of the proliferating stages of the parasite, with multiple applications in the genetic analysis of T. cruzi, including analyses of host-parasite interactions, gene expression regulation and drug development.