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1.
Proc Natl Acad Sci U S A ; 117(18): 9832-9839, 2020 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-32317383

RESUMO

G-quadruplex, assembled from a square array of guanine (G) molecules, is an important structure with crucial biological roles in vivo but also a versatile template for ordered functional materials. Although the understanding of G-quadruplex structures is the focus of numerous studies, little is known regarding the control of G-quartet stacking modes and the spontaneous orientation of G-quadruplex fibrils. Here, the effects of different metal ions and their concentrations on stacking modes of G-quartets are elucidated. Monovalent cations (typically K+) facilitate the formation of G-quadruplex hydrogels with both heteropolar and homopolar stacking modes, showing weak mechanical strength. In contrast, divalent metal ions (Ca2+, Sr2+, and Ba2+) at given concentrations can control G-quartet stacking modes and increase the mechanical rigidity of the resulting hydrogels through ionic bridge effects between divalent ions and borate. We show that for Ca2+ and Ba2+ at suitable concentrations, the assembly of G-quadruplexes results in the establishment of a mesoscopic chirality of the fibrils with a regular left-handed twist. Finally, we report the discovery of nematic tactoids self-assembled from G-quadruplex fibrils characterized by homeotropic fibril alignment with respect to the interface. We use the Frank-Oseen elastic energy and the Rapini-Papoular anisotropic surface energy to rationalize two different configurations of the tactoids. These results deepen our understanding of G-quadruplex structures and G-quadruplex fibrils, paving the way for their use in self-assembly and biomaterials.


Assuntos
DNA/química , Quadruplex G , Guanina/química , Hidrogéis/química , Anisotropia , Cátions Bivalentes/química , Cátions Monovalentes/química , DNA/ultraestrutura , Metabolismo Energético/efeitos dos fármacos , Líquidos Iônicos/química , Íons/química , Metais/química , Conformação de Ácido Nucleico/efeitos dos fármacos , Propriedades de Superfície
2.
Chemosphere ; 249: 126564, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32213389

RESUMO

Pb-based perovskite nanoparticles (PbPNPs) are amongst others used within highly efficient solar cells. PbPNPs can be released into the environment during their production, recycling or waste processing. In this study we investigated the fate and toxicity of PbPNPs on soil bacterial community under simulated natural environmental conditions across a range of pH, humic acid, and divalent cation concentrations. Increasing pH decreased PbPNPs-particle aggregation as well as Pb-ion release. The presence of only humic acid (HA) prevented the aggregation of PbPNPs-particles, whereas the presence of only divalent cations promoted the aggregation of PbPNPs-particles. The amount of Pb-ions released from the PbPNPs-particles was reduced in the presence of either HA or the divalent cations. Results of toxicity testing of PbPNPs by determining the metabolic potential of a bacterial community indicated that increasing pH alleviated particle toxicity. The presence of only HA reduced the toxicity of PbPNPs, while the presence of only divalent cations enhanced the particle toxicity. The coexistence of HA and divalent cations enhanced PbPNPs aggregation and reduced toxicity, with both Pb-ions and the interaction between the PbPNPs-particles and bacterial cells contributing to the toxic effects. Our study emphasized that environmental conditions play important roles that influencing the fate and toxicity of PbPNPs.


Assuntos
Bactérias/efeitos dos fármacos , Compostos de Cálcio/química , Nanopartículas/toxicidade , Óxidos/química , Microbiologia do Solo , Poluentes do Solo/toxicidade , Titânio/química , Cátions Bivalentes , Substâncias Húmicas/análise , Concentração de Íons de Hidrogênio , Íons , Chumbo , Solo , Poluentes do Solo/análise , Testes de Toxicidade
3.
Food Chem ; 320: 126623, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32220707

RESUMO

A novel ratiometric fluorescence probe was proposed for detecting lead (II) ions (Pb2+) in porphyra, the approach was based on copper nanoclusters and nitrogen-doped carbon quantum dots (CuNCs-CNQDs). In this probe, the CuNCs delivered the response signal, the fluorescence of which was enhanced by Pb2+ due to the aggregation-induced emission enhancement (AIEE) between Pb2+ and CuNCs. The CNQDs provided the self-calibration signal, whose fluorescence remained almost unchanged in coexistence with Pb2+. According to the change of fluorescence intensity ratio between the fluorophores, CuNCs-CNQDs nanohybrid was used as ratiometric probes for the sensitive detection of Pb2+ in the range of 0.010-2.5 mg L-1, with a detection limit of 0.0031 mg L-1. Finally, the probe was successfully applied to detect Pb2+ in porphyra with relative standard deviations (RSDs) lower than 5%. This study provides a straightforward, stable, and sensitive approach for detecting Pb2+ in porphyra.


Assuntos
Carbono/química , Cobre/química , Nitrogênio/química , Porphyra/química , Pontos Quânticos , Cátions Bivalentes/química , Fluorescência , Limite de Detecção , Nanoestruturas , Espectrometria de Fluorescência
4.
PLoS One ; 15(3): e0230327, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32150746

RESUMO

Sodium Calcium exchanger (NCX) proteins utilize the electrochemical gradient of Na+ to generate Ca2+ efflux (forward mode) or influx (reverse mode). In mammals, there are three unique NCX encoding genes-NCX1, NCX2, and NCX3, that comprise the SLC8A family, and mRNA from all three exchangers is expressed in hippocampal pyramidal cells. Furthermore, mutant ncx2-/- and ncx3-/- mice have each been shown to exhibit altered long-term potentiation (LTP) in the hippocampal CA1 region due to delayed Ca2+ clearance after depolarization that alters synaptic transmission. In addition to the role of NCX at the synapse of hippocampal subfields required for LTP, the three NCX isoforms have also been shown to localize to the dendrite of hippocampal pyramidal cells. In the case of NCX1, it has been shown to localize throughout the basal and apical dendrite of CA1 neurons where it helps compartmentalize Ca2+ between dendritic shafts and spines. Given the role for NCX and calcium in synaptic plasticity, the capacity of NCX splice-forms to influence backpropagating action potentials has clear consequences for the induction of spike-timing dependent synaptic plasticity (STDP). To explore this, we examined the effect of NCX localization, density, and allosteric activation on forward and back propagating signals and, next employed a STDP paradigm to monitor the effect of NCX on plasticity using back propagating action potentials paired with EPSPs. From our simulation studies we identified a role for the sodium calcium exchange current in normalizing STDP, and demonstrate that NCX is required at the postsynaptic site for this response. We also screened other mechanisms in our model and identified a role for the Ca2+ activated K+ current at the postsynapse in producing STDP responses. Together, our data reveal opposing roles for the Na+/Ca2+ exchanger current and the Ca2+ activated K+ current in setting STDP.


Assuntos
Região CA1 Hipocampal/fisiologia , Modelos Neurológicos , Plasticidade Neuronal/fisiologia , Células Piramidais/metabolismo , Trocador de Sódio e Cálcio/metabolismo , Potenciais de Ação/fisiologia , Animais , Região CA1 Hipocampal/citologia , Cálcio/metabolismo , Cátions Bivalentes/metabolismo , Cátions Monovalentes/metabolismo , Simulação por Computador , Modelos Animais , Potássio/metabolismo , Ratos , Sódio/metabolismo
5.
Environ Sci Technol ; 54(6): 3353-3362, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32083478

RESUMO

Although two-dimensional titanium carbide (Ti3C2Tx MXene) has emerged as a shining star material in various communities, its environmental behaviors and fate remain unknown. Herein, the colloidal properties and stability of Ti3C2Tx MXene are explored in aquatic systems for the first time, considering the roles of solution chemistry conditions (e.g., pH, ionic types, and strength). It was found that pH had no effect on the stability of Ti3C2Tx in the range of 5.0-11.0, whereas ionic valence and concentrations displayed significant effects on the aggregation behavior of Ti3C2Tx. By employing time-resolved dynamic light scattering measurements, the critical coagulation concentration (CCC) value of Ti3C2Tx was determined to be 12 mM for NaCl. The divalent cations Ca2+ and Mg2+ exhibited higher destabilizing capacity to Ti3C2Tx, as evidenced by the lower CCC values (0.3 and 0.4 mM for CaCl2 and MgCl2, respectively) and faster coagulation rates. Long-term stability studies implied that Ti3C2Tx MXene was less likely to be transported over long distances in the synthetic or natural waters. These findings provided significant insights into the fate and transport of Ti3C2Tx in the aquatic environment.


Assuntos
Cloreto de Sódio , Titânio , Cátions Bivalentes , Soluções
6.
Nat Commun ; 11(1): 922, 2020 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-32066742

RESUMO

Ryanodine receptors (RyR) are ion channels responsible for the release of Ca2+ from the sarco/endoplasmic reticulum and play a crucial role in the precise control of Ca2+ concentration in the cytosol. The detailed permeation mechanism of Ca2+ through RyR is still elusive. By using molecular dynamics simulations with a specially designed Ca2+ model, we show that multiple Ca2+ ions accumulate in the upper selectivity filter of RyR1, but only one Ca2+ can occupy and translocate in the narrow pore at a time, assisted by electrostatic repulsion from the Ca2+ within the upper selectivity filter. The Ca2+ is nearly fully hydrated with the first solvation shell intact during the whole permeation process. These results suggest a remote knock-on permeation mechanism and one-at-a-time occupation pattern for the hydrated Ca2+ within the narrow pore, uncovering the basis underlying the high permeability and low selectivity of the RyR channels.


Assuntos
Sinalização do Cálcio , Cálcio/metabolismo , Cátions Bivalentes/metabolismo , Simulação de Dinâmica Molecular , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Cálcio/química , Calpaína/metabolismo , Cátions Bivalentes/química , Canal de Liberação de Cálcio do Receptor de Rianodina/química , Eletricidade Estática , Relação Estrutura-Atividade
7.
Proc Natl Acad Sci U S A ; 117(10): 5291-5297, 2020 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-32098845

RESUMO

Heterotropic allosteric activation of protein function, in which binding of one ligand thermodynamically activates the binding of another, different ligand or substrate, is a fundamental control mechanism in metabolism and as such has been a long-aspired capability in protein design. Here we show that greatly increasing the magnitude of a protein's net charge using surface supercharging transforms that protein into an allosteric ligand- and counterion-gated conformational molecular switch. To demonstrate this we first modified the designed helical bundle hemoprotein H4, creating a highly charged protein which both unfolds reversibly at low ionic strength and undergoes the ligand-induced folding transition commonly observed in signal transduction by intrinsically disordered proteins in biology. As a result of the high surface-charge density, ligand binding to this protein is allosterically activated up to 1,300-fold by low concentrations of divalent cations and the polyamine spermine. To extend this process further using a natural protein, we similarly modified Escherichia coli cytochrome b 562 and the resulting protein behaves in a like manner. These simple model systems not only establish a set of general engineering principles which can be used to convert natural and designed soluble proteins into allosteric molecular switches useful in biodesign, sensing, and synthetic biology, the behavior we have demonstrated--functional activation of supercharged intrinsically disordered proteins by low concentrations of multivalent ions--may be a control mechanism utilized by Nature which has yet to be appreciated.


Assuntos
Grupo dos Citocromos b/química , Proteínas de Escherichia coli/química , Hemeproteínas/química , Proteínas Intrinsicamente Desordenadas/química , Engenharia de Proteínas/métodos , Regulação Alostérica , Cálcio/química , Cátions Bivalentes/química , Ligantes , Magnésio/química , Conformação Proteica , Dobramento de Proteína , Espermina/química , Termodinâmica
8.
Talanta ; 209: 120569, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31892050

RESUMO

As the concentration of Zn2+ in patients with prostate cancer is much less than that in healthy persons, Zn2+ concentration can be used as a marker to expediently screen prostate cancer. In this study, a sensitive and highly selective surface-enhanced Raman scattering (SERS) method to detect Zn2+ concentration in human prostatic fluids by utilizing water-insoluble 2-carboxyl-2'-hydroxyl-5'-sulfoformazylbenze (Zincon) as a SERS probe based on self-assembled Au nanoarrays at a liquid-liquid interface between n-hexane and Au colloids was proposed. Zincon showed remarkably different SERS bands before and after coordinating Zn2+ in the controlled conditions (70 µL of ethanol, 500 µL of n-hexane, pH value of 7.1 and 10 s of vortex mixing time), which can be used in quantifying Zn2+ with characteristic peaks. The proposed SERS method presented a good linear relationship ranging from 0.5 to 10 µmol/L and a satisfactory detection limit of 0.1 µmol/L as well as low interference with other metal ions. Moreover, the detection results are close to those of the conventional standard atomic absorption spectroscopy (AAS) method.


Assuntos
Ouro/química , Nanoestruturas/química , Neoplasias da Próstata/diagnóstico , Análise Espectral Raman/métodos , Zinco/análise , Cátions Bivalentes/análise , Formazans/química , Humanos , Limite de Detecção , Masculino , Nanoestruturas/ultraestrutura , Próstata/química , Próstata/patologia , Neoplasias da Próstata/química , Neoplasias da Próstata/patologia
9.
RNA ; 26(2): 111-125, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31776179

RESUMO

We develop an L-platform/L-scaffold framework we hypothesize may serve as a blueprint to facilitate site-specific RNA-cleaving nucleic acid enzyme design. Building on the L-platform motif originally described by Suslov and coworkers, we identify new critical scaffolding elements required to anchor a conserved general base guanine ("L-anchor") and bind functionally important metal ions at the active site ("L-pocket"). Molecular simulations, together with a broad range of experimental structural and functional data, connect the L-platform/L-scaffold elements to necessary and sufficient conditions for catalytic activity. We demonstrate that the L-platform/L-scaffold framework is common to five of the nine currently known naturally occurring ribozyme classes (Twr, HPr, VSr, HHr, Psr), and intriguingly from a design perspective, the framework also appears in an artificially engineered DNAzyme (8-17dz). The flexibility of the L-platform/L-scaffold framework is illustrated on these systems, highlighting modularity and trends in the variety of known general acid moieties that are supported. These trends give rise to two distinct catalytic paradigms, building on the classifications proposed by Wilson and coworkers and named for the implicated general base and acid. The "G + A" paradigm (Twr, HPr, VSr) exclusively utilizes nucleobase residues for chemistry, and the "G + M + " paradigm (HHr, 8-17dz, Psr) involves structuring of the "L-pocket" metal ion binding site for recruitment of a divalent metal ion that plays an active role in the chemical steps of the reaction. Finally, the modularity of the L-platform/L-scaffold framework is illustrated in the VS ribozyme where the "L-pocket" assumes the functional role of the "L-anchor" element, highlighting a distinct mechanism, but one that is functionally linked with the hammerhead ribozyme.


Assuntos
Engenharia Genética , Motivos de Nucleotídeos/genética , RNA Catalítico/genética , Catálise , Domínio Catalítico , Cátions Bivalentes/metabolismo , RNA/genética , RNA Catalítico/metabolismo
10.
Water Environ Res ; 92(5): 677-688, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31633854

RESUMO

The effect of mono- and divalent cation concentrations on digested sludge dewatering characteristics was investigated in a full-scale study at the Harrisonburg-Rockingham Regional Sewer Authority, VA (HRRSA) and at an additional POTW (POTW B). At HRRSA, anaerobic digesters were fed with primary sludge, enhanced biological phosphorus removal (EBPR) WAS, and trucked-in DAF solids from a poultry processing facility, and digested sludge was dewatered by belt filter press. The POTW B digesters received EBPR WAS and primary sludge, and dewatering was by centrifuge. Visual MINTEQ 3.1 (Gustafsson, 2014, Visual MINTEQ 3.1. User's manual) was used to determine cation speciation and struvite, tricalcium phosphate, vivianite, and ferrous sulfide formation at a range of digestion conditions. Significant Mg and Ca complexation was predicted at all conditions evaluated, and the majority of the complexes were monovalent cations. Complexation reduced the divalent cation concentration on the order of 40%. The HRRSA and POTW B data both showed that dewatered cake total solids (TS) increased linearly with uncomplexed divalent cation concentration and that there was an increase of 1.0%-1.1% TS for every additional 1.0 meq/L of uncomplexed divalent cations. Because POTW B used centrifuge dewatering, this relationship could be independent of dewatering technique. Further, the relationship between uncomplexed divalent cation concentration and dewaterability suggests that the selectivity of exocellular polymeric substances cation exchange sites is much greater for divalent cations so that monovalent cations only bind to them when their concentrations are much higher than those of the uncomplexed divalent cations. Thus, monovalent cation concentration may not be important in anaerobic digestion or other high alkalinity environments. PRACTITIONER POINTS: It is important to account for complexation in high alkalinity environments such as anaerobic digesters; The findings of this study suggest that dewatering characteristics of anaerobically digested enhanced biological phosphorus removal (EBPR) waste sludge can be significantly improved by preserving cation-mediated exocellular polymeric substances bonding in the solids stream by reducing struvite formation; Data from two full-scale POTW's indicated that dewatered cake TS was increased by 1.0%-1.1% for every 1.0 meq/L of dissolved uncomplexed divalent cations in the anaerobic digestate bulk solution; This relationship existed for dewatering both by a belt filter press and by a centrifuge suggesting that this relationship is independent of dewatering technology; Dissolved monovalent cation concentrations may not be important in anaerobic digestion or other high alkalinity environments unless their concentrations are much greater than those of the dissolved divalent cations.


Assuntos
Fósforo , Esgotos , Cátions Bivalentes , Estruvita , Eliminação de Resíduos Líquidos
11.
J Biosci Bioeng ; 129(2): 223-228, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31492609

RESUMO

Mercury pollution has always been a huge threat to human health due to its significant toxicity. Thus, it's the continuing goal to obtain new mercury detection techniques that are cost-effective, operational stable, performance efficient, and applicable to the environmental and biological milieus. In this research, the soluble pigment pyocyanin with anti-bacterial and anti-fungal activities, the biosynthesis pathway of which was engineered under the regulation of Hg2+-dependent transcriptional activator MerR, was firstly used as the visual detection signal in the whole-cell biosensor. The engineered biosensor displayed optical sensing window and a good linearity for Hg2+ in the range of 25-1000 nM, and the detection limit could reach as low as 10 nM. It permitted on-site detection of bioavailable Hg2+ with extraordinary selectivity and could resist the interferences of extra metal ions. What's more, the developed biosensor performed function well in a wide pH range (pH 4-10) as well as the environmental water. By fully imitating and utilizing the biosystems from nature, the engineered colorimetric biosensor has great economic and performance advantages over most chemosensors as well as whole-cell biosensors in the practical application of detecting Hg2+ in the contaminated aquatic systems.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Mercúrio/análise , Pseudomonas aeruginosa/metabolismo , Piocianina/biossíntese , Proteínas de Bactérias/genética , Técnicas Biossensoriais/métodos , Cátions Bivalentes/química , Colorimetria , Proteínas de Ligação a DNA/genética , Limite de Detecção , Mercúrio/química , Pseudomonas aeruginosa/genética
12.
Nucleic Acids Res ; 48(1): 332-348, 2020 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-31777930

RESUMO

Single-strand specific endoribonuclease YbeY has been shown to play an important role in the processing of the 3' end of the 16S rRNA in Escherichia coli. Lack of YbeY results in the accumulation of the 17S rRNA precursor. In contrast to a previous report, we show that Sinorhizobium meliloti YbeY exhibits endoribonuclease activity on single-stranded RNA substrate but not on the double-stranded substrate. This study also identifies the previously unknown metal ion involved in YbeY function to be Zn2+ and shows that the activity of YbeY is enhanced when the occupancy of zinc is increased. We have identified a pre-16S rRNA precursor that accumulates in the S. meliloti ΔybeY strain. We also show that ΔybeY mutant of Brucella abortus, a mammalian pathogen, also accumulates a similar pre-16S rRNA. The pre-16S species is longer in alpha-proteobacteria than in gamma-proteobacteria. We demonstrate that the YbeY from E. coli and S. meliloti can reciprocally complement the rRNA processing defect in a ΔybeY mutant of the other organism. These results establish YbeY as a zinc-dependent single-strand specific endoribonuclease that functions in 16S rRNA processing in both alpha- and gamma-proteobacteria.


Assuntos
Proteínas de Escherichia coli/genética , Escherichia coli/genética , Metaloproteínas/genética , RNA Ribossômico 16S/genética , Sinorhizobium meliloti/genética , Zinco/metabolismo , Pareamento de Bases , Cátions Bivalentes , Ensaios Enzimáticos , Escherichia coli/metabolismo , Expressão Gênica , Teste de Complementação Genética , Isoenzimas/deficiência , Isoenzimas/genética , Metaloproteínas/deficiência , Mutação , Conformação de Ácido Nucleico , Precursores de RNA/genética , Precursores de RNA/metabolismo , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/metabolismo , RNA Ribossômico 16S/metabolismo , Sinorhizobium meliloti/metabolismo
13.
Food Chem ; 306: 125578, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31622835

RESUMO

Oleosomes are storage vehicles of TAGs in plant seeds. They are protected with a phospholipid-protein monolayer and extracted with alkaline aqueous media; however, pH adjustment intensifies the extraction process. Therefore, the aim of this work was to investigate the extraction mechanism of rapeseed oleosomes at pH 7 and at the presence of monovalent and divalent cations (Na+, K+, Mg2+, and Ca+2). The oleosome yield at pH 9.5 was 64 wt%, while the yield at pH 7 with H2O was just 43 wt.%. The presence of cations at pH 7, significantly enhanced the yield, with K+ giving the highest yield (64 wt.%). The cations affected the oleosome interface and their interactions. The presence of monovalent cations resulted in aggregation and minor coalescence, while divalent cations resulted in extensive coalescence. These results help to understand the interactions of oleosomes in their native matrix and design simple extraction processes at neutral conditions.


Assuntos
Brassica/química , Cálcio/química , Magnésio/química , Extratos Vegetais/química , Potássio/química , Sódio/química , Cátions Bivalentes/química , Cátions Monovalentes/química , Concentração de Íons de Hidrogênio , Gotículas Lipídicas , Sementes/química , Água
14.
Nucleic Acids Res ; 48(3): 1406-1422, 2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-31863586

RESUMO

Recent advances in gene editing have been enabled by programmable nucleases such as transcription activator-like effector nucleases (TALENs) and CRISPR-Cas9. However, several open questions remain regarding the molecular machinery in these systems, including fundamental search and binding behavior as well as role of off-target binding and specificity. In order to achieve efficient and specific cleavage at target sites, a high degree of target site discrimination must be demonstrated for gene editing applications. In this work, we studied the binding affinity and specificity for a series of TALE proteins under a variety of solution conditions using in vitro fluorescence methods and molecular dynamics (MD) simulations. Remarkably, we identified that TALEs demonstrate high sequence specificity only upon addition of small amounts of certain divalent cations (Mg2+, Ca2+). However, under purely monovalent salt conditions (K+, Na+), TALEs bind to specific and non-specific DNA with nearly equal affinity. Divalent cations preferentially bind to DNA over monovalent cations, which attenuates non-specific interactions between TALEs and DNA and further stabilizes specific interactions. Overall, these results uncover new mechanistic insights into the binding action of TALEs and further provide potential avenues for engineering and application of TALE- or TALEN-based systems for genome editing and regulation.


Assuntos
Cálcio/química , Cátions Bivalentes/química , DNA/química , Magnésio/química , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/química , Sistemas CRISPR-Cas/genética , Proteínas de Ligação a DNA/química , Edição de Genes , Potássio/química , Ligação Proteica , Sódio/química , Soluções/química , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/metabolismo
15.
Protein Expr Purif ; 166: 105519, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31629955

RESUMO

Chitosanase (EC 3.2.1.132) is an important chitosan-degrading enzyme involved in industrial applications. In this study, a chitosanase gene (BbCSN-1) from Beauveria bassiana, an insect fungal pathogen, was cloned and expressed in Pichia pastoris. The amount of BbCSN-1 in the fermentation broth of P. pastoris gradually increased after induction with methanol from one to 6 d, reaching 398 µg/ml on the 6th day. The molecular characteristics of BbCSN-1 were measured with colloidal chitosan as a substrate. The purified BbCSN-1 exhibited optimum activity at pH 5 and 30 °C and was stable at pH 2-8 and below 40 °C. The Km value of BbCSN-1 was approximately 0.8 mg/ml at 30 °C (pH 6.0). The activity of BbCSN-1 was significantly enhanced by Mn2+ but inhibited by Co2+ and Cu2+. These results indicated that BbCSN-1 from B. bassiana could be easily expressed in P. pastoris, which provided a basis for further study on its application.


Assuntos
Beauveria/genética , Glicosídeo Hidrolases/genética , Pichia/genética , Proteínas Recombinantes/genética , Sequência de Aminoácidos , Cátions Bivalentes/química , Clonagem Molecular , Cobalto/química , Cobre/química , Expressão Gênica , Glicosídeo Hidrolases/química , Concentração de Íons de Hidrogênio , Manganês/química , Pichia/enzimologia , Ligação Proteica , Estabilidade Proteica , Proteínas Recombinantes/química , Temperatura , Termodinâmica
16.
Food Chem ; 303: 125374, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31473457

RESUMO

The application of a novel Pb2+ selective sensor based on the ionic imprinted polymer in determining trace levels of lead in natural water and in fruit juice was investigated using differential pulse voltammetry (DPV). The new and selective magnetic ion-imprinted polymer (Fe3O4@SiO2@IIP) was synthesized using 2-(2-aminophenyl)-1H-benzimidazole and 4-vinyl pyridine as a ligand and functional monomer, respectively. The new polymer was utilized for modifying a GCE to generate a new electrochemical sensor. Box-Behnken design (BBD) was utilized for optimizing the parameters influencing the peak current. Extremely suitable voltammetric performance was demonstrated for the electrochemical detection of lead with a low detection limit (0.05 ng mL-1), over a wide linear concentration range (0.1-80 ng mL-1). This modified electrode as a novel sensor was utilized for determining Pb(II) ions in natural water and in fruit juice with very reasonable outcomes.


Assuntos
Técnicas Eletroquímicas/métodos , Chumbo/análise , Polímeros/química , Cátions Bivalentes , Eletrodos , Compostos de Ferro/química , Chumbo/química , Limite de Detecção , Magnetismo , Dióxido de Silício/química
17.
Biosens Bioelectron ; 148: 111666, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31698301

RESUMO

As an essential transition-metal ion in organism, zinc ion (Zn2+) plays a significant role in cellular metabolism, apoptosis and neurotransmission. Herein, we reported a new intramolecular charge transfer-based two-photon probe, (E)-1-(6-(4-(2-(2-(2-methoxyethoxy)ethoxy)eth-oxy)-3,5-dimethylstyryl)quinolin-2-yl)-N,N-bis(pyridin-2-ylmethyl)methanamine (TEO-MPVQ), for ratiometric imaging and biosensing of Zn2+ in living cells and larval zebrafish. The maximum two-photon absorption cross-section values (δmax) of the developed probe increased from 115 ±â€¯16 GM to 188 ±â€¯27 GM, after the probe was chelated with Zn2+. The emission spectrum of TEO-MPVQ probe increased and red-shifted about 60 nm, resulting in the ratiometric determination of Zn2+ with high sensitivity and selectivity. The present fluorescent probe also demonstrated good biocompatibility, high spatial resolution and deep penetration depth for Zn2+ detection. Accordingly, TEO-MPVQ was able to track the endogenous Zn2+ release in neurons and also successfully applied for larval zebrafish imaging. By using of this effective probe, it was found that the endogenous Zn2+ was rapidly released from metallothionein under the stimulation of NO in neurons.


Assuntos
Técnicas Biossensoriais/métodos , Corantes Fluorescentes/química , Neurônios/química , Imagem Óptica/métodos , Peixe-Zebra , Zinco/análise , Animais , Cátions Bivalentes/análise , Larva/química , Metilaminas/química , Camundongos , Microscopia de Fluorescência por Excitação Multifotônica/métodos , Neurônios/citologia , Quinolinas/química , Espectrometria de Fluorescência/métodos , Peixe-Zebra/metabolismo
18.
J Environ Sci (China) ; 86: 78-86, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31787192

RESUMO

Permanganate treatment is widely used for disinfection of bacteria in surface-contaminated water. In this paper, the fate of the dissolved permanganate in aqueous solution after contact with cells of Pseudomonas fluorescens was studied. Concomitant accumulation of divalent cations of Mg2+, Zn2+, and Co2+ during precipitation of Mn oxides was also studied. The time course of the Mn concentration in solution showed an abrupt decrease after contact of Mn(VII) with microbial cells, followed by an increase after ~24 hr. XRD analysis of the precipitated Mn oxides, called biomass Mn oxides, showed the formation of low-crystalline birnessite. Visible spectroscopy and X-ray absorption near edge structure (XANES) analyses indicated that dissolved Mn(VII) was reduced to form biomass Mn oxides involving Mn(IV) and Mn(III), followed by reduction to soluble Mn(II). The numbers of electron transferred from microbial cells to permanganate and to biomass Mn oxides for 24 hr after the contact indicated that the numbers of electron transfer from microbial cell was approximately 50 times higher to dissolved permanganate than to the biomass Mn oxides in present experimental conditions. The 24 hr accumulation of divalent cations during formation of biomass Mn oxides was in the order of Co2+ > Zn2+ > Mg2+. XANES analysis of Co showed that oxidation of Co2+ to Co3+ resulted in higher accumulation of Co than Zn and Mg. Thus, treatment of surface water by KMnO4 solution is effective not only for disinfection of microorganisms, but also for the elimination of metal cations from surface water.


Assuntos
Biodegradação Ambiental , Compostos de Manganês/metabolismo , Óxidos/metabolismo , Microbiologia da Água , Poluentes Químicos da Água/metabolismo , Cátions Bivalentes , Cobalto/metabolismo , Magnésio/metabolismo , Poluentes Químicos da Água/análise , Zinco/metabolismo
19.
Phys Rev Lett ; 123(21): 218101, 2019 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-31809131

RESUMO

We develop an iterated map model to describe the bifurcations and complex dynamics caused by the feedback between voltage and intracellular Ca^{2+} and Na^{+} concentrations in paced ventricular myocytes. Voltage and Ca^{2+} can form either a positive or a negative feedback loop, while voltage and Na^{+} form a negative feedback loop. Under certain diseased conditions, when the feedback between voltage and Ca^{2+} is positive, Hopf bifurcations occur, leading to periodic oscillatory behaviors. When this feedback is negative, period-doubling bifurcation routes to alternans and chaos occur.


Assuntos
Cálcio/metabolismo , Modelos Cardiovasculares , Miócitos Cardíacos/metabolismo , Sódio/metabolismo , Relógios Biológicos , Cátions Bivalentes/metabolismo , Cátions Monovalentes/metabolismo , Membrana Celular/metabolismo , Polaridade Celular , Retroalimentação Fisiológica , Ventrículos do Coração/citologia , Ventrículos do Coração/metabolismo , Potenciais da Membrana , Miócitos Cardíacos/citologia , Trocador de Sódio e Cálcio/metabolismo
20.
Int J Mol Sci ; 20(23)2019 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-31795357

RESUMO

The present work focuses on unraveling the collisional processes leading to the fragmentation of the gas-phase furan molecules under the He+ and He2+ cations impact in the energy range 5-2000 eV. The presence of different mechanisms was identified by the analysis of the optical fragmentation spectra measured using the collision-induced emission spectroscopy (CIES) in conjunction with the ab initio calculations. The measurements of the fragmentation spectra of furan were performed at the different kinetic energies of both cations. In consequence, several excited products were identified by their luminescence. Among them, the emission of helium atoms excited to the 1s4d 1D2, 3D1,2,3 states was recorded. The structure of the furan molecule lacks an He atom. Therefore, observation of its emission lines is spectroscopic evidence of an impact reaction occurring via relocation of the electronic charge between interacting entities. Moreover, the recorded spectra revealed significant variations of relative band intensities of the products along with the change of the projectile charge and its velocity. In particular, at lower velocities of He+, the relative cross-sections of dissociation products have prominent resonance-like maxima. In order to elucidate the experimental results, the calculations have been performed by using a high level of quantum chemistry methods. The calculations showed that in both impact systems two collisional processes preceded fragmentation. The first one is an electron transfer from furan molecules to cations that leads to the neutralization and further excitation of the cations. The second mechanism starts from the formation of the He-C4H4O+/2+ temporary clusters before decomposition, and it is responsible for the appearance of the narrow resonances in the relative cross-section curves.


Assuntos
Furanos/química , Hélio/química , Cátions Bivalentes/química , Espectrometria de Massas , Modelos Moleculares , Temperatura
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