RESUMO
BACKGROUND: The Antarctic continent is a source of extreme microorganisms. Millions of years of isolation have produced unique biodiversity with adaptive responses to its extreme environment. Although the Antarctic climate is mainly cold, the presence of several geothermal sites, including thermal springs, fumaroles, hot soils and hydrothermal vents, provides ideal environments for the development of thermophilic and hyperthermophilic microorganisms. Their enzymes, called thermoenzymes, are the focus of interest in both academic and industrial research, mainly due to their high thermal activity and stability. Glutamate dehydrogenase, is an enzyme that plays a key role in the metabolism of carbon and nitrogen catalyzing reversibly the oxidative deamination of glutamate to alpha-ketoglutarate and ammonium. It belongs to the family of oxidoreductases, is widely distributed and it has been highly regarded for use as biosensors, particularly for their specificity and ability to operate in photochemical and electrochemical systems. However, the use of enzymes as biosensors is relatively problematic due to their instability to high temperatures, organic solvents and denaturing agents. The purpose of this study is to present the partial characterization of a thermophilic microorganism isolated from Deception Island, Antarctica, that displays glutamate dehydrogenase activity. RESULTS: In this work, we report the isolation of a thermophilic microorganism called PID15 from samples of Deception Island collected during the Antarctic Scientific Expedition ECA 46. This microorganism is a thermophile that grows optimally at 50 °C and pH 8.0. Scanning electron microscopy shows rod cells of 2.0 to 8.0 µm of length. Phylogenetic analysis of 16S rRNA gene revealed that this microorganism is closely related to Bacillus gelatini. This microorganism contains a thermostable glutamate dehydrogenase with optimal activity at pH 8.0 and temperatures for its activity from 37 to 50 °C, range of temperature of interest for biotechnological applications. This glutamate dehydrogenase is a highly thermostable enzyme. CONCLUSION: This is the first report of a microorganism from Antarctica containing a thermostable glutamate dehydrogenase that maintains its activity in a broad range of temperatures making it of potential interest for biotechnological applications.
Assuntos
Bactérias/enzimologia , Extremófilos/enzimologia , Glutamato Desidrogenase/análise , Animais , Regiões Antárticas , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Extremófilos/genética , Extremófilos/crescimento & desenvolvimento , Ilhas , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Fatores de TempoRESUMO
BACKGROUND: The Antarctic continent is a source of extreme microorganisms. Millions of years of isolation have produced unique biodiversity with adaptive responses to its extreme environment. Although the Antarctic climate is mainly cold, the presence of several geothermal sites, including thermal springs, fumaroles, hot soils and hydrothermal vents, provides ideal environments for the development of thermophilic and hyperthermophilic microorganisms. Their enzymes, called thermoenzymes, are the focus of interest in both academic and industrial research, mainly due to their high thermal activity and stability. Glutamate dehydrogenase, is an enzyme that plays a key role in the metabolism of carbon and nitrogen catalyzing reversibly the oxidative deamination of glutamate to alpha-ketoglutarate and ammonium. It belongs to the family of oxidoreductases, is widely distributed and it has been highly regarded for use as biosensors, particularly for their specificity and ability to operate in photochemical and electrochemical systems. However, the use of enzymes as biosensors is relatively problematic due to their instability to high temperatures, organic solvents and denaturing agents. The purpose of this study is to present the partial characterization of a thermophilic microorganism isolated from Deception Island, Antarctica, that displays glutamate dehydrogenase activity. RESULTS: In this work, we report the isolation of a thermophilic microorganism called PID15 from samples of Deception Island collected during the Antarctic Scientific Expedition ECA 46. This microorganism is a thermophile that grows optimally at 50 °C and pH 8.0. Scanning electron microscopy shows rod cells of 2.0 to 8.0 µm of length. Phylogenetic analysis of 16S rRNA gene revealed that this microorganism is closely related to Bacillus gelatini. This microorganism contains a thermostable glutamate dehydrogenase with optimal activity at pH 8.0 and temperatures for its activity from 37 to 50 °C, range of temperature of interest for biotechnological applications. This glutamate dehydrogenase is a highly thermostable enzyme. CONCLUSION: This is the first report of a microorganism from Antarctica containing a thermostable glutamate dehydrogenase that maintains its activity in a broad range of temperatures making it of potential interest for biotechnological applications.
Assuntos
Animais , Bactérias/enzimologia , Extremófilos/enzimologia , Glutamato Desidrogenase/análise , Filogenia , Fatores de Tempo , Bactérias/crescimento & desenvolvimento , Bactérias/genética , RNA Ribossômico 16S/genética , Microscopia Eletrônica de Varredura , Reação em Cadeia da Polimerase , Microscopia Eletrônica de Transmissão , Ilhas , Extremófilos/crescimento & desenvolvimento , Extremófilos/genética , Regiões AntárticasRESUMO
Four lipases were purified from ID17, a thermophilic bacterium belonging to Geobacillus genus isolated from Deception Island, Antarctica. Lipase activity was detected by opacity test and p-nitrophenyl laurate methods. Lipase production was better in a medium containing tryptone as the carbon and nitrogen source, without non-ionic detergents and pH 7.5. Proteins were ultrafiltered from supernatant and separated using anion exchange and size exclusion chromatography resulting in four distinct fractions with lipase activity (called Lip1-4). Purified lipases showed an optimal pH at 9.0, 9.5, 10.0 and 8.0 and temperature at 65, 70, 75 and 80 °C for Lip1-4, respectively. Lip1 and Lip2 showed higher activity using p-nitrophenol decanoate as substrate, whereas Lip3 and Lip4 prefer p-nitrophenol laurate. Based on their molecular weight Lip1 and Lip2 are trimeric and pentameric proteins, respectively, whereas Lip3 and Lip4 are monomeric proteins. Lip1 was exceptionally thermostable maintaining 70 % of its activity after incubating it at 70 °C for 8 h. Based on their characteristics, the four lipases obtained from ID17 are good candidates to understand the mechanisms of lipase stability and to be used in different types of industrial applications.
Assuntos
Geobacillus/enzimologia , Lipase/isolamento & purificação , Lipase/metabolismo , Regiões Antárticas , Geobacillus/química , Concentração de Íons de Hidrogênio , Lauratos/metabolismo , Lipase/química , Nitrofenóis/metabolismo , Multimerização Proteica , Especificidade por SubstratoRESUMO
In Antarctica microorganisms are exposed to several conditions that trigger the generation of reactive oxygen species, such as high UV radiation. Under these conditions they must have an important antioxidant defense system in order to prevent oxidative damage. One of these defenses are pigments which are part of the non-enzymatic antioxidant mechanisms. In this work we focused on the antioxidant capacity of pigments from an Antarctic microorganism belonging to Pedobacter genus. This microorganism produces different types of pigments which belong to the carotenoids group. The antioxidant capacity of a mix of pigments was analyzed by three different methods: 1,1-diphenyl-2-picrylhydrazyl, ROS detection and oxygen electrode. The results obtained from these approaches indicate that the mix of pigments has a strong antioxidant capacity. The oxidative damage induced by UVB exposure to liposomes was also analyzed. Intercalated pigments within the liposomes improved its resistance to lipid peroxidation. Based on the analysis carried out along this research we conclude that the antioxidant properties of the mix of pigments protect this bacterium against oxidative damage. These properties make this mix of pigments a powerful antioxidant mixture with potential biotechnological applications.