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Characterizing the mycobacterial transporters involved in the uptake and/or catabolism of host-derived nutrients required by mycobacteria may identify novel drug targets against tuberculosis. Here, we identify and characterize a member of the amino acid-polyamine-organocation superfamily, a potential γ-aminobutyric acid (GABA) transport protein, GabP, from Mycobacterium smegmatis The protein was expressed to a level allowing its purification to homogeneity, and size exclusion chromatography coupled with multiangle laser light scattering (SEC-MALLS) analysis of the purified protein showed that it was dimeric. We showed that GabP transported γ-aminobutyric acid both in vitro and when overexpressed in E. coli Additionally, transport was greatly reduced in the presence of ß-alanine, suggesting it could be either a substrate or inhibitor of GabP. Using GabP reconstituted into proteoliposomes, we demonstrated that γ-aminobutyric acid uptake is driven by the sodium gradient and is stimulated by membrane potential. Molecular docking showed that γ-aminobutyric acid binds MsGabP, another Mycobacterium smegmatis putative GabP, and the Mycobacterium tuberculosis homologue in the same manner. This study represents the first expression, purification, and characterization of an active γ-aminobutyric acid transport protein from mycobacteria.IMPORTANCE The spread of multidrug-resistant tuberculosis increases its global health impact in humans. As there is transmission both to and from animals, the spread of the disease also increases its effects in a broad range of animal species. Identifying new mycobacterial transporters will enhance our understanding of mycobacterial physiology and, furthermore, provides new drug targets. Our target protein is the gene product of msmeg_6196, annotated as GABA permease, from Mycobacterium smegmatis strain MC2 155. Our current study demonstrates it is a sodium-dependent GABA transporter that may also transport ß-alanine. As GABA may well be an essential nutrient for mycobacterial metabolism inside the host, this could be an attractive target for the development of new drugs against tuberculosis.
Asunto(s)
Proteínas Bacterianas/metabolismo , Transporte Biológico/fisiología , Proteínas Transportadoras de GABA en la Membrana Plasmática/metabolismo , Mycobacterium smegmatis/metabolismo , Transportadores de Anión Orgánico/metabolismo , Sodio/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Proteínas Bacterianas/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli , Proteínas Transportadoras de GABA en la Membrana Plasmática/genética , Regulación Bacteriana de la Expresión Génica , Metabolómica , Simulación del Acoplamiento Molecular , Transportadores de Anión Orgánico/genética , Filogenia , Ácido gamma-Aminobutírico/química , Ácido gamma-Aminobutírico/genéticaRESUMEN
Biochar has become a popular soil amendment. However, its effect on soil microbial community is still unclear. In the present study, maize straw biochar was pyrolysed at 300°C, 450°C and 600°C, respectively, and then was added to agricultural soil at the ratio of 0.5%, 1% and 2%. Bacterial dynamics was analyzed in the pot experiments using denaturing gradient gel electrophoresis. The results indicated that the pyrolysis temperature has great impact on the elemental composition, pH and porous structures of biochar. Moreover, pyrolysis temperature was primary factor to drive the variation of bacterial community structure in biochar amended soil. In addition, the results suggested that biochar amendments on agricultural soil would decrease the bacterial diversity, and selectively promote growth of functional bacteria to become the dominant community, which could increase the bacterial community organization and improve the stability of bacteria to counteract effects of perturbation.
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Carbón Orgánico/química , Microbiota , Microbiología del Suelo , Suelo/química , Zea mays/química , Agricultura/métodos , Bacterias/crecimiento & desarrolloRESUMEN
Glutamate transporters are responsible for uptake of the neurotransmitter glutamate in mammalian central nervous systems. Their archaeal homologue GltPh, an aspartate transporter isolated from Pyrococcus horikoshii, has been the focus of extensive studies through crystallography, MD simulations and single-molecule FRET (smFRET). Here, we summarize the recent research progress on GltPh, in the hope of gaining some insights into the transport mechanism of this aspartate transporter.
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Sistema de Transporte de Aminoácidos X-AG/metabolismo , Pyrococcus horikoshii/metabolismo , Ácido Aspártico/metabolismo , Transporte Biológico , Conformación Proteica , Especificidad por SustratoRESUMEN
Geobacillus thermodenitrificans NG80-2 encodes two long-chain NAD(P)-dependent alcohol dehydrogenases, gtADH1 and gtADH2, in the terminal oxidation pathway of long-chain n-alkanes for the conversion of long-chain alkyl alcohols to their corresponding aldehydes. Both gtADH1 and gtADH2 are thermostable enzymes and oxidize long-chain alkyl alcohols up to at least C(30). In order to understand the structural basis for their role in long-chain alkane degradation, we have crystallized gtADH2. Single, colourless crystals were obtained from a recombinant preparation of ADH2 overexpressed in Escherichia coli. The crystals belong to space group C222(1), with unit-cell parameters a = 56.0, b = 99.6, c = 123.1 Å. Diffraction data were collected in-house to 1.79 Å resolution. The crystals contain one monomer in the asymmetric unit, with a V(M) value of 2.17 Å(3) Da(-1) and an estimated solvent content of 43%.
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Oxidorreductasas de Alcohol/química , Geobacillus/enzimología , NADP/metabolismo , Cromatografía en Gel , Cristalización , Cristalografía por Rayos X , Multimerización de Proteína , SolucionesRESUMEN
Li-rich Mn-based oxides (LRMOs) are considered as one of the most-promising cathode materials for next generation Li-ion batteries (LIBs) because of their high energy density. Nevertheless, the intrinsic shortcomings, such as the low first coulomb efficiency, severe capacity/voltage fade, and poor rate performance seriously limit its commercial application in the future. In this work, we construct successfully g-C3N4 coating layer to modify Li1.2Mn0.54Ni0.13Co0.13O2 (LMNC) via a facile solution. The g-C3N4 layer can alleviate the side-reaction between electrolyte and LMNC materials, and improve electronic conduction of LMNC. In addition, the g-C3N4 layer can suppress the collapse of structure and improve cyclic stability of LMNC materials. Consequently, g-C3N4 (4 wt%)-coated LMNC sample shows the highest initial coulomb efficiency (78.5%), the highest capacity retention ratio (78.8%) and the slightest voltage decay (0.48 V) after 300 loops. Besides, it also can provide high reversible capacity of about 300 and 93 mAh g-1 at 0.1 and 10C, respectively. This work proposes a novel approach to achieve next-generation high-energy density cathode materials, and g-C3N4 (4 wt%)-coated LMNC shows an enormous potential as the cathode materials for next generation LIBs with excellent performance.
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ZnTiO3 and ZnTiO3-CeO2 microspheres with particle sizes of about 100-300 nm were synthesized for the first time by a simple solvothermal process followed by calcination. The results indicate that CeO2 modification does not alter the morphology of the microspheres. ZnTiO3-CeO2 (0, 3, 6, and 9 wt%) show an initial charge (discharge) capacity of 171.01 (253.2), 204.6 (507.5), 213.4 (451.6) and 126.2 (367.2) mA h g-1 at 500 mA g-1, respectively. After 500 cycles, the corresponding charge (discharge) capacities were 191.1 (192.3), 298.7 (300.3), 322.4 (328.5) and 211.2 (212.3) mA h g-1, respectively. Obviously, the charge (discharge) capacities of the ZnTiO3-CeO2 composites are superior to those of pristine ZTO, which demonstrates that the Li storage performance of the CeO2-modified ZTO electrodes is improved. The CeO2 shell provides a good electronic contact between ZnTiO3 and CeO2, decreasing charge transfer resistance and facilitating the charge transportation of the ZnTiO3-CeO2 composite. In addition, the formed phase interface between CeO2 and ZnTiO3 may provide more active sites for electrochemical reactions, improving the reversibility of Li-ion intercalation and decreasing the electrochemical polarization during cycling, especially at high current densities. Therefore, such ZnTiO3-CeO2 microspheres can be regarded as hopeful candidates for anode materials for Li-ion batteries.
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Bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed in aquatic environments. To date, limited knowledge is available on their temporal and spatial variations in freshwater environments. Here an investigation of HNA and LNA bacterial abundance and their flow cytometric characteristics was conducted in an exorheic river (Haihe River, Northern China) over a one year period covering September (autumn) 2011, December (winter) 2011, April (spring) 2012, and July (summer) 2012. The results showed that LNA and HNA bacteria contributed similarly to the total bacterial abundance on both the spatial and temporal scale. The variability of HNA on abundance, fluorescence intensity (FL1) and side scatter (SSC) were more sensitive to environmental factors than that of LNA bacteria. Meanwhile, the relative distance of SSC between HNA and LNA was more variable than that of FL1. Multivariate analysis further demonstrated that the influence of geographical distance (reflected by the salinity gradient along river to ocean) and temporal changes (as temperature variation due to seasonal succession) on the patterns of LNA and HNA were stronger than the effects of nutrient conditions. Furthermore, the results demonstrated that the distribution of LNA and HNA bacteria, including the abundance, FL1 and SSC, was controlled by different variables. The results suggested that LNA and HNA bacteria might play different ecological roles in the exorheic river.
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Bacterias/aislamiento & purificación , ADN Bacteriano/análisis , Ácidos Nucleicos/análisis , Ríos/microbiología , Microbiología del Agua , China , Monitoreo del Ambiente/métodos , Citometría de Flujo , Agua Dulce , Geografía , Concentración de Iones de Hidrógeno , Análisis Multivariante , Estaciones del Año , Análisis Espacio-Temporal , TemperaturaRESUMEN
Environmental microbes utilize four degradation pathways for the oxidation of n-alkanes. Although the enzymes degrading n-alkanes in different microbes may vary, enzymes functioning in the first step in the aerobic degradation of alkanes all belong to the alkane hydroxylases. Alkane hydroxylases are a class of enzymes that insert oxygen atoms derived from molecular oxygen into different sites of the alkane terminus (or termini) depending on the type of enzymes. In this review, we summarize the different types of alkane hydroxylases, their degrading steps, and compare typical enzymes from various classes with regard to their three-dimensional structures, in order to provide insights into how the enzymes mediate their different roles in the degradation of n-alkanes and what determines their different substrate ranges. Through the above analyzes, the degrading mechanisms of enzymes can be elucidated and molecular biological methods can be utilized to expand their catalytic roles in the petrochemical industry or in bioremediation of oil-contaminated environments.