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1.
Chemosphere ; 254: 126905, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32957298

RESUMO

With the development of modern technologies, the exploitation and application of rare earth metals (REMs) have increased parallelly. Consequently, more REMs are entering into the environment and therefore there is a pressing need to assess their potential environmental hazards. Here, a standard toxicity test with wheat (Triticum aestivum) was conducted to investigate the single and mixture toxicity of La and Ce in solutions with different levels of calcium and nitrilotriacetic acid (NTA) and results were deciphered by different modeling approaches. Both La and Ce caused adverse effect to wheat, but the presence of Ca and NTA alleviated their toxicity. The obtained EC50 for [La] or [Ce] changed by more than 28-fold and by 4-fold, respectively, with the increase of Ca or NTA. The biotic ligand model (BLM) explained approximately 93% variation of single La or Ce toxicity. The binding constants obtained were 4.14, 6.67, and 6.59 for logKCaBL, logKLaBL, and logKCeBL respectively. The electrostatic toxicity model (ETM) was proved as effective as the BLM, with R2 = 0.93 for La and R2 = 0.92 for Ce. For La-Ce mixtures, parameters from single toxicity approaches were applied successfully to predict the mixture toxicity with concentration addition (CA) model based on the BLM or ETM theory (R2 = 0.92 and RMSE = 8.56; R2 = 0.90 and RMSE = 9.6, respectively). Thus, the results obtained in this study prove that both ETM and BLM theories are appropriate to predict single and mixture REMs toxicity, providing coherent and promising tools for the risk assessment of REM pollution.


Assuntos
Cálcio/química , Cério/toxicidade , Lantânio/toxicidade , Ácido Nitrilotriacético/química , Testes de Toxicidade/métodos , Triticum/efeitos dos fármacos , Ligantes , Modelos Teóricos , Soluções , Eletricidade Estática , Triticum/crescimento & desenvolvimento
2.
Nat Commun ; 11(1): 3881, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32753572

RESUMO

Cells typically respond to chemical or physical perturbations via complex signaling cascades which can simultaneously affect multiple physiological parameters, such as membrane voltage, calcium, pH, and redox potential. Protein-based fluorescent sensors can report many of these parameters, but spectral overlap prevents more than ~4 modalities from being recorded in parallel. Here we introduce the technique, MOSAIC, Multiplexed Optical Sensors in Arrayed Islands of Cells, where patterning of fluorescent sensor-encoding lentiviral vectors with a microarray printer enables parallel recording of multiple modalities. We demonstrate simultaneous recordings from 20 sensors in parallel in human embryonic kidney (HEK293) cells and in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), and we describe responses to metabolic and pharmacological perturbations. Together, these results show that MOSAIC can provide rich multi-modal data on complex physiological responses in multiple cell types.


Assuntos
Técnicas Biossensoriais/métodos , Células-Tronco Pluripotentes Induzidas/metabolismo , Microscopia de Fluorescência/métodos , Miócitos Cardíacos/metabolismo , Imagem Óptica/métodos , Potenciais de Ação/efeitos dos fármacos , Antagonistas Adrenérgicos beta/farmacologia , Técnicas Biossensoriais/instrumentação , Cálcio/química , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Peróxido de Hidrogênio/farmacologia , Concentração de Íons de Hidrogênio , Células-Tronco Pluripotentes Induzidas/citologia , Mitocôndrias/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/fisiologia , Imagem Óptica/instrumentação , Oxidantes/farmacologia , Oxirredução/efeitos dos fármacos , Propanolaminas/farmacologia
3.
Nature ; 583(7814): 66-71, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32612224

RESUMO

Dental enamel is a principal component of teeth1, and has evolved to bear large chewing forces, resist mechanical fatigue and withstand wear over decades2. Functional impairment and loss of dental enamel, caused by developmental defects or tooth decay (caries), affect health and quality of life, with associated costs to society3. Although the past decade has seen progress in our understanding of enamel formation (amelogenesis) and the functional properties of mature enamel, attempts to repair lesions in this material or to synthesize it in vitro have had limited success4-6. This is partly due to the highly hierarchical structure of enamel and additional complexities arising from chemical gradients7-9. Here we show, using atomic-scale quantitative imaging and correlative spectroscopies, that the nanoscale crystallites of hydroxylapatite (Ca5(PO4)3(OH)), which are the fundamental building blocks of enamel, comprise two nanometric layers enriched in magnesium flanking a core rich in sodium, fluoride and carbonate ions; this sandwich core is surrounded by a shell with lower concentration of substitutional defects. A mechanical model based on density functional theory calculations and X-ray diffraction data predicts that residual stresses arise because of the chemical gradients, in agreement with preferential dissolution of the crystallite core in acidic media. Furthermore, stresses may affect the mechanical resilience of enamel. The two additional layers of hierarchy suggest a possible new model for biological control over crystal growth during amelogenesis, and hint at implications for the preservation of biomarkers during tooth development.


Assuntos
Amelogênese , Esmalte Dentário/química , Ácidos/química , Cálcio/química , Carbonatos/química , Cristalização , Teoria da Densidade Funcional , Esmalte Dentário/ultraestrutura , Durapatita/química , Fluoretos/química , Humanos , Magnésio/química , Microscopia Eletrônica de Transmissão e Varredura , Sódio/química , Tomografia , Difração de Raios X
4.
PLoS Comput Biol ; 16(7): e1007996, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32667909

RESUMO

Cortical spreading depression (CSD) is the propagation of a relatively slow wave in cortical brain tissue that is linked to a number of pathological conditions such as stroke and migraine. Most of the existing literature investigates the dynamics of short term phenomena such as the depolarization and repolarization of membrane potentials or large ion shifts. Here, we focus on the clinically-relevant hour-long state of neurovascular malfunction in the wake of CSDs. This dysfunctional state involves widespread vasoconstriction and a general disruption of neurovascular coupling. We demonstrate, using a mathematical model, that dissolution of calcium that has aggregated within the mitochondria of vascular smooth muscle cells can drive an hour-long disruption. We model the rate of calcium clearance as well as the dynamical implications on overall blood flow. Based on reaction stoichiometry, we quantify a possible impact of calcium phosphate dissolution on the maintenance of F0F1-ATP synthase activity.


Assuntos
Depressão Alastrante da Atividade Elétrica Cortical , Potenciais da Membrana , Mitocôndrias/metabolismo , Vasoconstrição , Trifosfato de Adenosina/química , Cálcio/química , Fosfatos de Cálcio/química , Córtex Cerebral/fisiopatologia , Circulação Cerebrovascular , Citosol/química , Retículo Endoplasmático/química , Substância Cinzenta/fisiopatologia , Humanos , Modelos Teóricos , Acoplamento Neurovascular , Oscilometria , Oxigênio/química , Fosforilação , ATPases Translocadoras de Prótons/química , Acidente Vascular Cerebral/fisiopatologia
5.
Food Chem ; 333: 127493, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-32659659

RESUMO

The effects of the addition of salt ions and molecular weights (Mw) of CH on Mesona chinensis polysaccharide (MCP)-chitosan (CH) hydrogel were investigated. Result indicated both low concentration of monovalent salt ions (Na+ and K+), divalent cations (Ca2+) and oxoanions (SO42-) could promote the gel properties of MCP-CH hydrogel. The Mw of CH has huge impact on the formation and properties of hydrogel. Combining the relationship between rheology and structural, monovalent salt ions such as Na+ and K+ affect gel formation and its properties by influencing electrostatic interaction and chain conformation. Both divalent cations (Ca2+) and oxoanions (SO42-) facilitated the formation of gel networks via electrostatic interaction, coordination bonds and hydrogen bonds. Moreover, Mw of CH influenced formation and texture of MCP-CH hydrogel via affecting the conformation of CH molecular chain. These findings will provide a few theoretical bases to understand the formation mechanism of MCP-CH hydrogel.


Assuntos
Quitosana/química , Hidrogéis/química , Lamiaceae/química , Extratos Vegetais/química , Polieletrólitos/química , Sais/química , Ânions/química , Cálcio/química , Hidrogéis/síntese química , Peso Molecular , Polissacarídeos/química , Potássio/química , Reologia , Sódio/química
6.
Food Chem ; 332: 127440, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32652411

RESUMO

Rehydration in an alkaline solution has been shown to improve the rehydration behaviour of milk protein isolate (MPI). In this study, the focus is on citric acid neutralization of MPI powder dissolved in alkaline solution. The results showed that alkalization induced more negative zeta-potential compared to MPI control, reducing it from -22.4 mV to -32.6 mV. Neutralization had a relatively similar zeta-potential value as alkalized sample. Sodium carbonate addition increased pH and caused a consequential reduction of ionic calcium in aqueous phase and, neutralization caused a small increase in ionic calcium. Soluble aggregate of κ-casein protein and whey protein was suggested in alkalization and neutralization process by non-reducing SDS-PAGE. In addition, neutralization kept a stable colloidal particle size for pHs decreased to pH 9,8 and 7; however, alkalization and neutralization created casein aggregates of larger colloidal particle size than primary casein micelle in control MPI.


Assuntos
Cálcio/química , Caseínas/química , Quelantes/química , Animais , Bovinos , Ácido Cítrico/química , Concentração de Íons de Hidrogênio , Micelas , Tamanho da Partícula , Proteínas do Soro do Leite/química
7.
Proc Natl Acad Sci U S A ; 117(31): 18470-18476, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690682

RESUMO

Lipid membrane fusion is an essential process for a number of critical biological functions. The overall process is thermodynamically favorable but faces multiple kinetic barriers along the way. Inspired by nature's engineered proteins such as SNAP receptor [soluble N-ethylmale-imide-sensitive factor-attachment protein receptor (SNARE)] complexes or viral fusogenic proteins that actively promote the development of membrane proximity, nucleation of a stalk, and triggered expansion of the fusion pore, here we introduce a synthetic fusogen that can modulate membrane fusion and equivalently prime lipid membranes for calcium-triggered fusion. Our fusogen consists of a gold nanoparticle functionalized with an amphiphilic monolayer of alkanethiol ligands that had previously been shown to fuse with lipid bilayers. While previous efforts to develop synthetic fusogens have only replicated the initial steps of the fusion cascade, we use molecular simulations and complementary experimental techniques to demonstrate that these nanoparticles can induce the formation of a lipid stalk and also drive its expansion into a fusion pore upon the addition of excess calcium. These results have important implications in general understanding of stimuli-triggered fusion and the development of synthetic fusogens for biomedical applications.


Assuntos
Cálcio/metabolismo , Membrana Celular/metabolismo , Ouro/química , Bicamadas Lipídicas/metabolismo , Nanopartículas Metálicas/química , Cálcio/química , Membrana Celular/química , Ouro/metabolismo , Humanos , Bicamadas Lipídicas/química , Fusão de Membrana , Simulação de Dinâmica Molecular , Proteínas SNARE/metabolismo , Análise Serial de Tecidos
8.
Proc Natl Acad Sci U S A ; 117(31): 18504-18510, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32699145

RESUMO

The human blood protein vitronectin (Vn) is a major component of the abnormal deposits associated with age-related macular degeneration, Alzheimer's disease, and many other age-related disorders. Its accumulation with lipids and hydroxyapatite (HAP) has been demonstrated, but the precise mechanism for deposit formation remains unknown. Using a combination of solution and solid-state NMR experiments, cosedimentation assays, differential scanning fluorimetry (DSF), and binding energy calculations, we demonstrate that Vn is capable of binding both soluble ionic calcium and crystalline HAP, with high affinity and chemical specificity. Calcium ions bind preferentially at an external site, at the top of the hemopexin-like (HX) domain, with a group of four Asp carboxylate groups. The same external site is also implicated in HAP binding. Moreover, Vn acquires thermal stability upon association with either calcium ions or crystalline HAP. The data point to a mechanism whereby Vn plays an active role in orchestrating calcified deposit formation. They provide a platform for understanding the pathogenesis of macular degeneration and other related degenerative disorders, and the normal functions of Vn, especially those related to bone resorption.


Assuntos
Cálcio/metabolismo , Durapatita/metabolismo , Degeneração Macular/metabolismo , Vitronectina/metabolismo , Sítios de Ligação , Cálcio/química , Durapatita/química , Humanos , Ligação Proteica , Vitronectina/química
9.
Chemosphere ; 260: 127624, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32683029

RESUMO

Soil organic matter (SOM) is known to exert a major control on the mobility and bioavailability of cationic nutrients. However, the role of SOM in the fate of anionic nutrients, especially phosphorus (P), is less well characterized. The objectives of this study were to (1) compare the formation of binary complexes of calcium (Ca) with humic acids (HA) extracted from two contrasting soils, and (2) determine if binary HA-Ca complexes could incorporate P by forming ternary HA-Ca-P complexes. The Ca binding capacities of the HA extracted from an agricultural organic soil (AOS) and a pristine riparian soil (RS) were measured via potentiometric titrations; the formation of ternary complexes was analyzed by size fractionation using MWCO tubes. Proton and Ca binding capacities of RS-HA were higher than AOS-HA, and pH had a weaker effect on Ca binding to RS-HA. These differences are consistent with lower proportions of aromatic groups, and a higher proportion of alkyl groups derived from 13C NMR spectroscopy. Together, the NMR, titration and MWCO data indicate that Ca binds to RS-HA through monodentate complexes and electrostatic attraction that are capable of binding P producing ternary complexes. In contrast, at pH 8.5 Ca forms bidentate complexes with AOS-HA, which do not provide bridging positions to incorporate P. Overall, our results imply that the formation of HA-Ca and HA-Ca-P complexes depend on the structure of the HA, and that complexation to HA may play an important role in the fate of P in terrestrial and aquatic environments.


Assuntos
Cálcio/química , Fósforo/análise , Poluentes do Solo/análise , Ânions , Cálcio na Dieta , Cátions , Substâncias Húmicas/análise , Minerais , Solo/química , Poluentes do Solo/química
10.
Nat Commun ; 11(1): 3068, 2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32555155

RESUMO

Surgical adhesions are bands of scar tissues that abnormally conjoin organ surfaces. Adhesions are a major cause of post-operative and dialysis-related complications, yet their patho-mechanism remains elusive, and prevention agents in clinical trials have thus far failed to achieve efficacy. Here, we uncover the adhesion initiation mechanism by coating beads with human mesothelial cells that normally line organ surfaces, and viewing them under adhesion stimuli. We document expansive membrane protrusions from mesothelia that tether beads with massive accompanying adherence forces. Membrane protrusions precede matrix deposition, and can transmit adhesion stimuli to healthy surfaces. We identify cytoskeletal effectors and calcium signaling as molecular triggers that initiate surgical adhesions. A single, localized dose targeting these early germinal events completely prevented adhesions in a preclinical mouse model, and in human assays. Our findings classifies the adhesion pathology as originating from mesothelial membrane bridges and offer a radically new therapeutic approach to treat adhesions.


Assuntos
Cálcio/química , Epitélio/metabolismo , Aderências Teciduais/metabolismo , Animais , Sinalização do Cálcio , Adesão Celular , Linhagem Celular , Membrana Celular/metabolismo , Biologia Computacional , Citoesqueleto/metabolismo , Citosol/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Imageamento Tridimensional , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Complicações Pós-Operatórias , Análise de Componente Principal , RNA Interferente Pequeno/metabolismo , Análise de Célula Única
11.
Chemosphere ; 256: 127021, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32438128

RESUMO

The effect of gamma-irradiation doses of 0, 10, 100, and 500 kGy at the dose rates of 1 or 0.1 kGy/h on the molecular and chemical properties of humic substances (HS) were investigated using total organic carbon measurements, UV-Vis spectrometry, 13C nuclear magnetic resonance, and acid-base titration. A possible mechanism of the radiolysis on HS was also speculated. The complexation ability of irradiated HS with Ca2+ ions was studied using a Ca ion-selective electrode. The apparent formation constants of the Ca-HS complexes increased as the irradiation dose increased, and this was attributed to the relative increase in the ratio of phenolic -OH to carboxylic groups of HS. The contribution of the phenolic -OH groups to Ca-HS complexes was suppressed at pH 5 owing to its high acid dissociation constants. In addition, the radiation dose rates of 1 and 0.1 kGy/h did not significantly affect the properties of HS and the apparent formation constants of the Ca-HS complexes.


Assuntos
Cálcio/química , Raios gama , Substâncias Húmicas , Íons
12.
Proc Natl Acad Sci U S A ; 117(21): 11217-11219, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32393617

RESUMO

Understanding the timing and mechanisms of amino acid synthesis and racemization on asteroidal parent bodies is key to demonstrating how amino acids evolved to be mostly left-handed in living organisms on Earth. It has been postulated that racemization can occur rapidly dependent on several factors, including the pH of the aqueous solution. Here, we conduct nanoscale geochemical analysis of a framboidal magnetite grain within the Tagish Lake carbonaceous chondrite to demonstrate that the interlocking crystal arrangement formed within a sodium-rich, alkaline fluid environment. Notably, we report on the discovery of Na-enriched subgrain boundaries and nanometer-scale Ca and Mg layers surrounding individual framboids. These interstitial coatings would yield a surface charge state of zero in more-alkaline fluids and prevent assimilation of the individual framboids into a single grain. This basic solution would support rapid synthesis and racemization rates on the order of years, suggesting that the low abundances of amino acids in Tagish Lake cannot be ascribed to fluid chemistry.


Assuntos
Aminoácidos , Meteoroides , Sódio/química , Água/química , Aminoácidos/biossíntese , Aminoácidos/síntese química , Colúmbia Britânica , Cálcio/química , Óxido Ferroso-Férrico/química , Concentração de Íons de Hidrogênio , Lagos , Magnésio/química , Estereoisomerismo , Tomografia/métodos
13.
Chemosphere ; 250: 126245, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32234617

RESUMO

Hydrothermal experiments with magnesium, ammonium, and phosphate (MAP) solution at a temperature of 120 OC for 24 h and pH (9 and 10), whilst effects of varying Mg2+/Ca2+/HCO-3 ratios on struvite crystallization were examined. The study was performed to investigate their effects on the quality and quantity of crystals using the XRPD Rietveld refinement and SEM method. Obviously, the struvite crystallization was inhibited through the forming of calcite, dolomite, hydroxyapatite, sylvite, and Mg-whitlockite under different pH conditions. In the absence of Ca2+ and HCO-3 ions, struvite and dittmarite were formed at pH solutions (9 and 10). Struvite proportion reduced with pH (9 and 10) under Mg2+/Ca2+/HCO-3 ratios (1:1:1 and 2:1:1), and depleted under the Mg2+/Ca2+/HCO-3 ratio of 1:2:2. An obvious change in morphologies of crystals into nanosized particles was observed. Results of the low proportion of struvite for experiments with Mg2+/Ca2+/HCO-3 molar ratios may be a drawback for phosphate recovery.


Assuntos
Cálcio/química , Estruvita/química , Compostos de Amônio , Carbonato de Cálcio , Cálcio na Dieta , Cristalização , Íons , Magnésio , Compostos de Magnésio/química , Fosfatos/química
14.
Food Chem ; 321: 126680, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32247181

RESUMO

Seed phytic acid reduces mineral bioavailability by chelating minerals. Consumption of common bean seeds with the low phytic acid 1 (lpa1) mutation improved iron status in human trials but caused adverse gastrointestinal effects, presumably due to increased stability of lectin phytohemagglutinin L (PHA-L) compared to the wild type (wt). A hard-to-cook (HTC) defect observed in lpa1 seeds intensified this problem. We quantified the HTC phenotype of lpa1 common beans with three genetic backgrounds. The HTC phenotype in the lpa1 black bean line correlated with the redistribution of calcium particularly in the cell walls, providing support for the "phytase-phytate-pectin" theory of the HTC mechanism. Furthermore, the excess of free cations in the lpa1 mutation in combination with different PHA alleles affected the stability of PHA-L lectin.


Assuntos
Cálcio/química , Lectinas/química , Phaseolus/química , Ácido Fítico/química , Fito-Hemaglutininas/química , Culinária , Dureza , Temperatura Alta , Mutação , Phaseolus/genética , Sementes/química , Sementes/genética
15.
Food Chem ; 322: 126709, 2020 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-32283376

RESUMO

This study elucidated the formation mechanism of calcium-indigo carmine (IC) complex. Parameters of the complex, such as Turbiscan stability index (TSI), yield and stoichiometric ratio (Ca to IC) were investigated. Isothermal titration calorimetry (ITC), UV/Visible spectrophotometry, infrared spectroscopy and Raman spectroscopy were used to explore the interaction between IC and Ca(II) during formation reaction and in the powdered lake. The TSI analysis and yield test suggested pH 7.0 as the most suitable pH to produce this complex. The conjugation between IC and Ca(II) was an exothermal reaction with increase of entropy (ΔH = -0.728 ± 0.11 kJ·mol-1 and ΔS = 66.55 ± 1.89 kJ·mol-1·K-1 at 25 °C). Both ITC and stoichiometric ratio test suggested one Ca atom tended to conjugate five IC molecules at ambient temperature when an adequate amount of Ca(II) appeared. Spectrometry techniques suggested that the sulfonic acid groups in IC formed coordination bonds with the Ca to construct the complex.


Assuntos
Cálcio/química , Corantes de Alimentos/química , Índigo Carmim/química , Calorimetria , Cor , Entropia , Espectrofotometria , Espectrofotometria Infravermelho
16.
PLoS Comput Biol ; 16(4): e1007791, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32282806

RESUMO

Widefield calcium imaging enables recording of large-scale neural activity across the mouse dorsal cortex. In order to examine the relationship of these neural signals to the resulting behavior, it is critical to demix the recordings into meaningful spatial and temporal components that can be mapped onto well-defined brain regions. However, no current tools satisfactorily extract the activity of the different brain regions in individual mice in a data-driven manner, while taking into account mouse-specific and preparation-specific differences. Here, we introduce Localized semi-Nonnegative Matrix Factorization (LocaNMF), a method that efficiently decomposes widefield video data and allows us to directly compare activity across multiple mice by outputting mouse-specific localized functional regions that are significantly more interpretable than more traditional decomposition techniques. Moreover, it provides a natural subspace to directly compare correlation maps and neural dynamics across different behaviors, mice, and experimental conditions, and enables identification of task- and movement-related brain regions.


Assuntos
Algoritmos , Mapeamento Encefálico/métodos , Cálcio/metabolismo , Processamento de Imagem Assistida por Computador/métodos , Córtex Pré-Frontal/diagnóstico por imagem , Animais , Cálcio/química , Camundongos , Córtex Pré-Frontal/química
17.
Science ; 368(6489): 428-433, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32327598

RESUMO

The melanocortin-4 receptor (MC4R) is involved in energy homeostasis and is an important drug target for syndromic obesity. We report the structure of the antagonist SHU9119-bound human MC4R at 2.8-angstrom resolution. Ca2+ is identified as a cofactor that is complexed with residues from both the receptor and peptide ligand. Extracellular Ca2+ increases the affinity and potency of the endogenous agonist α-melanocyte-stimulating hormone at the MC4R by 37- and 600-fold, respectively. The ability of the MC4R crystallized construct to couple to ion channel Kir7.1, while lacking cyclic adenosine monophosphate stimulation, highlights a heterotrimeric GTP-binding protein (G protein)-independent mechanism for this signaling modality. MC4R is revealed as a structurally divergent G protein-coupled receptor (GPCR), with more similarity to lipidic GPCRs than to the homologous peptidic GPCRs.


Assuntos
Cálcio/química , Receptor Tipo 4 de Melanocortina/química , Receptores Acoplados a Proteínas-G/química , Cristalografia por Raios X , AMP Cíclico/química , Humanos , Ligantes , Hormônios Estimuladores de Melanócitos/química , Hormônios Estimuladores de Melanócitos/farmacologia , Mutação , Canais de Potássio Corretores do Fluxo de Internalização/química , Ligação Proteica , Multimerização Proteica , Estrutura Secundária de Proteína , Receptor Tipo 4 de Melanocortina/antagonistas & inibidores , Receptor Tipo 4 de Melanocortina/genética , Receptores Acoplados a Proteínas-G/antagonistas & inibidores , Receptores Acoplados a Proteínas-G/genética , Transdução de Sinais
18.
Nat Struct Mol Biol ; 27(4): 382-391, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32251414

RESUMO

The bestrophin family of calcium (Ca2+)-activated chloride (Cl-) channels, which mediate the influx and efflux of monovalent anions in response to the levels of intracellular Ca2+, comprises four members in mammals (bestrophin 1-4). Here we report cryo-EM structures of bovine bestrophin-2 (bBest2) bound and unbound by Ca2+ at 2.4- and 2.2-Å resolution, respectively. The bBest2 structure highlights four previously underappreciated pore-lining residues specifically conserved in Best2 but not in Best1, illustrating the differences between these paralogs. Structure-inspired electrophysiological analysis reveals that, although the channel is sensitive to Ca2+, it has substantial Ca2+-independent activity for Cl-, reflecting the opening at the cytoplasmic restriction of the ion conducting pathway even when Ca2+ is absent. Moreover, the ion selectivity of bBest2 is controlled by multiple residues, including those involved in gating.


Assuntos
Bestrofinas/ultraestrutura , Canais de Cloreto/ultraestrutura , Conformação Proteica , Animais , Bestrofinas/química , Bestrofinas/genética , Cálcio/química , Bovinos , Canais de Cloreto/química , Canais de Cloreto/genética , Microscopia Crioeletrônica , Citoplasma/química , Citoplasma/genética , Citoplasma/ultraestrutura , Humanos , Ativação do Canal Iônico/genética , Ligação Proteica/genética , Transdução de Sinais
19.
Food Chem ; 320: 126625, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32203839

RESUMO

The cellular antioxidant activity (CAA) assay is wildly used for quantifying antioxidant activities of foods and dietary supplements in vitro. Among various incubation and handling buffers used in different laboratories, the inconsistence in concentrations of ions, particularly calcium and magnesium, has somehow been neglected. We hired the Hank's balanced salt solution with or without calcium and magnesium to perform CAA assay in Caco-2 cells and HepG2 cells, evaluating the impacts of these cations. The absence of calcium and magnesium reduced intracellular ROS level and underestimated the CAA of quercetin, Trolox and catechin. The abnormally high extracellular calcium and magnesium can also produce inaccurate results. Hank's buffer is recommended to ensure the accuracy and reproducibility. It elucidates precautions must be taken on these cations' concentrations of the buffers while conducting CAA determinations on different types of cells and when comparing foods and beverages with various calcium/magnesium contents.


Assuntos
Antioxidantes/farmacologia , Cálcio/farmacologia , Magnésio/farmacologia , Cálcio/química , Linhagem Celular Tumoral , Humanos , Magnésio/química , Quercetina , Reprodutibilidade dos Testes
20.
Food Chem ; 317: 126412, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32113138

RESUMO

In this study, three whey protein concentrate systems enriched in α-lactalbumin, produced using membrane filtration (LAC-M), selective precipitation (LAC-P) and ion-exchange chromatography (LAC-IE), were fortified with calcium chloride (CaCl2) at 0-5 mM and changes in physicochemical properties studied. Binding of calcium (Ca2+) occurred for LAC-P in the range 0.00-2.00 mM, with an affinity constant (Kd) of 1.63 × 10-7, resulting in a proportion of total protein-bound calcium of 81.8% at 2 mM CaCl2. At 5 mM CaCl2, LAC-P had volume mean diameter (VMD) of 638 nm, while LAC-M and LAC-IE had VMD of 204 and 3.87 nm, respectively. Changes in physicochemical properties were dependent on the approach used to enrich α-lactalbumin and concentrations of other macromolecules (e.g., phospholipid). The results obtained in this study provide fundamental insights into the influence of fortification with soluble calcium salts on the physicochemical stability of next-generation whey protein ingredients enriched in α-lactalbumin.


Assuntos
Cálcio/química , Lactalbumina/química , Proteínas do Soro do Leite/química , Cálcio/metabolismo , Cloreto de Cálcio/química , Cálcio na Dieta , Cromatografia por Troca Iônica , Alimentos Fortificados , Tamanho da Partícula
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