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Water use has been increasing globally by 1% per year, and recycling and re-use are critical issues compromised by the presence of pollutants. In this context, the design of novel materials and/or procedures for the large scale-removal of pollutants must be economically and environmentally feasible in order to be considered as part of the solution by emerging economies. We demonstrate that the cross-linking of biodegradable polysaccharides such as starch, dextrin, or dextrin and ß-cyclodextrin with divinyl sulfone is an innovative strategy for synthesizing insoluble and eco-friendly sorbent polymers, including pSt, pDx and pCD-Dx. The evaluation of these polymers' ability to remove ciprofloxacin (CIP), a prime example of antibiotic pollution, revealed that pSt, with a Kd of 1469 L/kg and a removal rate higher than 92%, is a favorable material. Its sorption is pH-dependent and enhanced at a mildly alkaline pH, allowing for the desorption (i.e., cleaning) and reuse of pSt through an environmentally friendly treatment with 20 mM AcONa pH 4.6. The facts that pSt (i) shows a high affinity for CIP even at high NaCl concentrations, (ii) can be obtained from affordable starting materials, and (iii) is synthesized and regenerated through organic, solvent-free procedures make pSt a novel sustainable material for inland water and seawater remediation, especially in less developed countries, due to its simplicity and low cost.
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This article presents the outdoor and synthetic performance data concerning the main electrical parameters estimated from the I-V curve for three photovoltaic technologies (HIT, m-Si and CIGS) and the weather conditions (irradiance, ambient and panel temperature). Synthetic data were generated by simulating in OpenModelica software the impact of weather conditions on device performance, considering an irradiance range between 50 and 1300 W/m2. The outdoor data corresponds to the performance of the evaluated PV modules in outdoor tests in Medellin-Colombia for ten months using capacitive I-V tracers. In both cases, different capacitor values were considered to evaluate the effect on the I-V curve behavior of devices.
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Assessments of ecosystem service and function losses of wetlandscapes (i.e., wetlands and their hydrological catchments) suffer from knowledge gaps regarding impacts of ongoing hydro-climatic change. This study investigates hydro-climatic changes during 1976-2015 in 25 wetlandscapes distributed across the world's tropical, arid, temperate and cold climate zones. Results show that the wetlandscapes were subject to precipitation (P) and temperature (T) changes consistent with mean changes over the world's land area. However, arid and cold wetlandscapes experienced higher T increases than their respective climate zone. Also, average P decreased in arid and cold wetlandscapes, contrarily to P of arid and cold climate zones, suggesting that these wetlandscapes are located in regions of elevated climate pressures. For most wetlandscapes with available runoff (R) data, the decreases were larger in R than in P, which was attributed to aggravation of climate change impacts by enhanced evapotranspiration losses, e.g. caused by land-use changes.
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Ozone (O3) therapy has been used for medical procedures for centuries; however, there are no extensive studies on its utilization in horses. This study aimed to evaluate the application of transrectal O3 on horses by physical and laboratorial evaluation, and production of reactive oxygen species (ROS). Sixteen healthy horses were separated in two groups: a control group (CG) and a group treated with O3 (TG). The TG animals received 1L of an oxygen and O3 mixture transrectally. The initial dose was 10µg/ml for the first two applications, 15µg/ml for the following two applications, and 20µg/ml for the next six applications. The CG animals received 1L of oxygen transrectally. In TG animals no variations in the physical examination were detected; furthermore, TG animals did not exhibit changes in biochemical evaluation results, fibrinogen concentrations, or ROS production. TG animals had increased red blood cell counts, hemoglobin concentrations, and packet cell volume values in comparison to the baseline and CG values. We could infer that O3 affected the red blood cell counts and improved rhetological properties of the blood. The transrectal application of O3 in horses is safe and can indirectly improve the oxygenation and metabolism of tissues.(AU)
A utilização medicinal do ozônio (O3) é secular, contudo não existem estudos expressivos de sua utilização em equinos. O objetivo deste estudo foi avaliar o efeito da aplicação transretal de O3 em equinos por meio da avaliação física, laboratorial, e produção de espécies reativas de oxigênio (EROs). Dezesseis equinos hígidos foram separados em dois grupos: grupo controle (GC) e grupo tratado com O3 (GT). O GT recebeu por via retal 1L da mistura de oxigênio e ozônio, sendo a dose inicial de 10µg/ml por duas aplicações, 15µg/ml por mais duas aplicações e 20µg/ml por seis aplicações. O GC recebeu 1L de oxigênio via transretal. No GT não foram observadas alterações no exame físico, bem como não foram observadas alterações na avaliação bioquímica, concentração de fibrinogênio e produção de EROs. O GT apresentou aumento no número de hemácias, na concentração de hemoglobina, e nos valores de hematócrito em relação aos valores basais e GC. Podemos inferir que o O3 alterou os valores de eritrócitos e melhorou as propriedades reológicas do sangue. Conclui-se que a aplicação transretal de 03 em equinos é segura e pode melhorar indiretamente a oxigenação e metabolismo dos tecidos.(AU)
Assuntos
Animais , Ozônio/uso terapêutico , Administração Retal , Espécies Reativas de Oxigênio , Cavalos/sangue , AntioxidantesRESUMO
Saponins are potential wide-spectrum antitumor drugs, and copper(I) catalyzed azide-alkyne 1,3-dipolar cycloaddition is a suitable approach to synthesizing saponin-like compounds by regioselective glycosylation of the C2/C3 hydroxyl and C28 carboxylic groups of triterpene aglycones maslinic acid (MA) and oleanolic acid (OA). Biological studies on the T-84 human colon carcinoma cell line support the role of the hydroxyl groups at C2/C3, the influence of the aglycone, and the bulky nature of the substituents in C28. OA bearing a α-d-mannose moiety at C28 (compound 18) focused our interest because the estimated inhibitory concentration 50 was similar to that reported for ginsenoside Rh2 against colon cancer cells and it inhibits the G1-S phase transition affecting the cell viability and apoptosis. Considering that triterpenoids from natural sources have been identified as inhibitors of nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) signaling, docking studies were conducted to evaluate whether NF-κB may be a potential target. Results are consistent with the biological study and predict a similar binding mode of MA and compound 18 to the p52 subunit from NF-κB but not for OA. The fact that the binding site is shared by the NF-κB inhibitor 6,6-dimethyl-2-(phenylimino)-6,7-dihydrobenzo[d][1,3]oxathiol-4(5H)-one supports the result and points to NF-κB as a potential target of both MA and compound 18.
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There are various neoplasms and tumour-like conditions of the pleura. Mesothelioma is perhaps the most widely recognised; however, there are many others that are more common and should be considered. Understanding the similarities and differences can be helpful in managing the patient with a newly found pleural lesion. We will discuss clinical symptoms at presentation and describe the imaging findings associated with these tumours, starting with conventional radiology, and correlating with computed tomography and combined positron-emission tomography (PET)/computed tomography (CT). Finally, imaging characteristics that help differentiation between the benign and malignant varieties will be reviewed.
Assuntos
Neoplasias Pulmonares/diagnóstico por imagem , Mesotelioma/diagnóstico por imagem , Pleura/diagnóstico por imagem , Neoplasias Pleurais/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Tomografia Computadorizada por Raios X , Diagnóstico Diferencial , Fluordesoxiglucose F18 , Humanos , Neoplasias Pulmonares/patologia , Mesotelioma/patologia , Mesotelioma Maligno , Estadiamento de Neoplasias , Pleura/patologia , Neoplasias Pleurais/classificação , Neoplasias Pleurais/patologia , Compostos RadiofarmacêuticosRESUMO
Despite the importance of the adjuvant in the immunization process, very few adjuvants merge with the antigens in vaccines. A synthetic self-adjuvant oleic-vinyl sulfone (OVS) linked to the catalytic region of recombinant serine/threonine phosphatase 2A from the nematode Angiostrongylus costaricensis (rPP2A) was used for intranasal immunization in mice previously infected with Trichuris muris The animal intranasal immunization with rPP2A-OVS showed a reduction of 99.01% in the number of the nematode eggs and 97.90% in adult. The immunohistochemical analysis of the intestinal sections showed that in immunized animals with lipopeptide the mucus was significantly higher than in the other experimental groups. Also, these animals presented significantly different chemokine, CCL20 and CCL11, levels. However, although the number and size of Tuft cells did not vary between groups, the intensity of fluorescence per cell was significant in the group immunized with the rPP2A-OVS. The results of the present study suggest that mice immunized with the lipopeptide are capable of activating a combined Th17/Th9 response. This strategy of immunization may be of great applicability not only in immunotherapy and immunoprophylaxis to control diseases caused by nematodes but also in pathologies necessitating action at the level of the Th9 response in the intestinal mucosa.
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Adjuvantes Imunológicos/administração & dosagem , Proteínas de Helminto/administração & dosagem , Lipopeptídeos/administração & dosagem , Proteína Fosfatase 2/administração & dosagem , Sulfonas/administração & dosagem , Tricuríase/prevenção & controle , Vacinas Conjugadas/administração & dosagem , Adjuvantes Imunológicos/síntese química , Administração Intranasal , Sequência de Aminoácidos , Animais , Quimiocina CCL11/genética , Quimiocina CCL11/imunologia , Quimiocina CCL20/genética , Quimiocina CCL20/imunologia , Feminino , Expressão Gênica , Proteínas de Helminto/biossíntese , Proteínas de Helminto/imunologia , Interleucinas/genética , Interleucinas/imunologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/imunologia , Mucosa Intestinal/parasitologia , Lipopeptídeos/biossíntese , Lipopeptídeos/imunologia , Camundongos , Camundongos Endogâmicos AKR , Contagem de Ovos de Parasitas , Proteína Fosfatase 2/biossíntese , Proteína Fosfatase 2/imunologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/imunologia , Alinhamento de Sequência , Sulfonas/química , Sulfonas/imunologia , Células Th17/efeitos dos fármacos , Células Th17/imunologia , Células Th17/parasitologia , Tricuríase/imunologia , Tricuríase/parasitologia , Trichuris/efeitos dos fármacos , Trichuris/imunologiaRESUMO
BACKGROUND: Around 20%-30% of breast cancers overexpress the proto-oncogene human epidermal growth receptor 2 (HER2), and they are characterized by being very invasive. Therefore, many current studies are focused on testing new therapies against tumors that overexpress this receptor. In particular, there exists major interest in new strategies to fight breast cancer resistant to trastuzumab (Tmab), a humanized antibody that binds specifically to HER2 interfering with its mitogenic signaling. Our team has previously developed immunostimulating complexes (ISCOMs) as nanocapsules functionalized with lipid vinyl sulfones, which can incorporate protein A and bind to G immunoglobulins that makes them very flexible nanocarriers. METHODS AND RESULTS: The aim of this in vitro study was to synthesize and evaluate a drug delivery system based on protein A-functionalized ISCOMs to target HER2-overexpressing cells. We describe the preparation of ISCOMs, the loading with the drugs doxorubicin and paclitaxel, the binding of ISCOMs to alkyl vinyl sulfone-protein A, the coupling of Tmab, and the evaluation in both HER2-overexpressing breast cancer cells (HCC1954) and non-overexpressing cells (MCF-7) by flow cytometry and fluorescence microscopy. Results show that the uptake is dependent on the level of overexpression of HER2, and the analysis of the cell viability reveals that targeted drugs are selective toward HCC1954, whereas MCF-7 cells remain unaffected. CONCLUSION: Protein A-functionalized ISCOMs are versatile carriers that can be coupled to antibodies that act as targeting agents to deliver drugs. When coupling to Tmab and loading with paclitaxel or doxorubicin, they become efficient vehicles for the selective delivery of the drug to Tmab-resistant HER2-overexpressing breast cancer cells. These nanoparticles may pave the way for the development of novel therapies for poor prognosis resistant patients.
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Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Sistemas de Liberação de Medicamentos/métodos , ISCOMs/química , Lipídeos/química , Receptor ErbB-2/metabolismo , Sulfonas/química , Trastuzumab/uso terapêutico , Antineoplásicos/farmacologia , Neoplasias da Mama/patologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Endocitose/efeitos dos fármacos , Feminino , Citometria de Fluxo , Humanos , Células MCF-7 , Nanopartículas/química , Nanopartículas/ultraestrutura , Oxazinas/metabolismo , Paclitaxel/farmacologia , Paclitaxel/uso terapêutico , Proto-Oncogene Mas , Proteína Estafilocócica A/química , Trastuzumab/farmacologiaRESUMO
A novel one-pot method for the synthesis of polyethyleneimine (PEI)-coated gold nanoparticles (AuPEI-NPs) that combines the reductant-stabilizer properties of PEI with microwave irradiation starting from hydrogen tetrachloroaurate acid (HAuCl4 ) and branched PEI 25â kDa (b25kPEI) was explored. The method was straightforward, green, and low costing, for which the Au/PEI ratio (1:1 to 1:128â w/w) was a key parameter to modulate their capabilities as DNA delivery nanocarriers. Transfection assays in CHO-k1 cells demonstrated that AuPEI-NPs with 1:16 and 1:32â w/w ratios behaved as effective DNA gene vectors with improved transfection efficiencies (twofold) and significantly lower toxicity than unmodified b25kPEI and Lipofectamineâ 2000. The transfection mediated by these AuPEI-NP-DNA polyplexes preferentially used the caveolae-mediated route for intracellular internalization, as shown by studies performed by using specific internalization inhibitors as well as colocalization with markers of clathrin- and caveolae-dependent pathways. The AuPEI-NP polyplexes preferentially used the more efficient caveolae internalization pathway to promote transfection, a fact that supports their higher transfection efficiency relative to that of Lipofectamineâ 2000. In addition, intracellular trafficking of the AuPEI-NPs was studied by transmission electron microscopy.
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DNA/metabolismo , Portadores de Fármacos/química , Ouro/química , Nanopartículas Metálicas/química , Polietilenoimina/química , Animais , Células CHO , Cavéolas/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Cricetinae , Cricetulus , DNA/química , Portadores de Fármacos/toxicidade , Difusão Dinâmica da Luz , Vetores Genéticos/metabolismo , Nanopartículas Metálicas/toxicidade , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , RNA Interferente Pequeno/química , RNA Interferente Pequeno/metabolismo , Espectrofotometria Ultravioleta , TransfecçãoRESUMO
Gene transfection mediated by the cationic polymer polyethylenimine (PEI) is considered a standard methodology. However, while highly branched PEIs form smaller polyplexes with DNA that exhibit high transfection efficiencies, they have significant cell toxicity. Conversely, low molecular weight PEIs (LMW-PEIs) with favorable cytotoxicity profiles display minimum transfection activities as a result of inadequate DNA complexation and protection. To solve this paradox, a novel polyelectrolyte complex was prepared by the ionic cross-linking of branched 1.8 kDa PEI with citric acid (CA). This system synergistically exploits the good cytotoxicity profile exhibited by LMW-PEI with the high transfection efficiencies shown by highly branched and high molecular weight PEIs. The polyectrolyte complex (1.8 kDa-PEI@CA) was obtained by a simple synthetic protocol based on the microwave irradiation of a solution of 1.8 kDa PEI and CA. Upon complexation with DNA, intrinsic properties of the resulting particles (size and surface charge) were measured and their ability to form stable polyplexes was determined. Compared with unmodified PEIs the new complexes behave as efficient gene vectors and showed enhanced DNA binding capability associated with facilitated intracellular DNA release and enhanced DNA protection from endonuclease degradation. In addition, while transfection values for LMW-PEIs are almost null, transfection efficiencies of the new reagent range from 2.5- to 3.8-fold to those of Lipofectamine 2000 and 25 kDa PEI in several cell lines in culture such as CHO-k1, FTO2B hepatomas, L6 myoblasts, or NRK cells, simultaneously showing a negligible toxicity. Furthermore, the 1.8 kDa-PEI@CA polyelectrolyte complexes retained the capability to transfect eukaryotic cells in the presence of serum and exhibited the capability to promote in vivo transfection in mouse (as an animal model) with an enhanced efficiency compared to 25 kDa PEI. Results support the polyelectrolyte complex of LMW-PEI and CA as promising generic nonviral gene carriers.
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Ácido Cítrico/química , Eletrólitos/química , Polietilenoimina/química , Transfecção , Linhagem Celular , Humanos , Técnicas In Vitro , Peso MolecularRESUMO
AIM: The aim of this study was to describe a case series of patients with primary or secondary antiphospholipid syndrome (APS) treated with direct oral anticoagulants (DOACs). PATIENTS AND METHODS: Clinical charts of eight patients with thrombotic primary or secondary APS treated with direct oral anticoagulants (DOACs) between January 2012 and May 2015 were reviewed. RESULTS: The mean age was 45 ± 14.36 (range 27-69 years). Four patients had secondary APS (50%). All patients were initially treated with warfarin by a mean time of 70.87 ± 57.32 months (range 17-153 months). Changes in anticoagulation were defined by recurring thrombosis in five patients (62.5%) and life-threatening bleeding in the other three cases. Seven patients (87.5%) received rivaroxaban treatment and one patient (12.5%) apixaban. The mean follow-up period with DOACs was 19 ± 10.06 months (range 2-36 months). There was no recurrence of thrombosis by the time of data collection. CONCLUSIONS: Despite not being the standard treatment in APS, we propose DOACs as a rational alternative for the management of patients with this diagnosis. Further interventional clinical studies are necessary for possible standardization of this therapy in APS patients.
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Anticoagulantes/uso terapêutico , Síndrome Antifosfolipídica/tratamento farmacológico , Hemorragia/induzido quimicamente , Trombose/prevenção & controle , Administração Oral , Adulto , Idoso , Anticoagulantes/administração & dosagem , Anticoagulantes/efeitos adversos , Síndrome Antifosfolipídica/complicações , Feminino , Seguimentos , Hemorragia/epidemiologia , Humanos , Pessoa de Meia-Idade , Pirazóis/efeitos adversos , Pirazóis/uso terapêutico , Piridonas/efeitos adversos , Piridonas/uso terapêutico , Recidiva , Estudos Retrospectivos , Rivaroxabana/efeitos adversos , Rivaroxabana/uso terapêutico , Trombose/etiologia , Resultado do Tratamento , Varfarina/efeitos adversos , Varfarina/uso terapêuticoRESUMO
The receptor for advanced glycation end products (RAGE) is involved in diabetes or angiogenesis in tumors. Under pathological conditions, RAGE is overexpressed and upon ligand binding and internalization stimulates signaling pathways that promote cell proliferation. In this work, amino dendritic polymers PEI 25 kDa and alkylated derivatives of PAMAM-G2 were engineered by the nonenzymatic Maillard glycation reaction to generate novel AGE-containing gene delivery vectors targeting the RAGE. The glycated dendritic polymers were easily prepared and retained the capability to bind and protect DNA from endonucleases. Furthermore, while glycation decreased the transfection efficiency of the dendriplexes in CHO-k1 cells which do not express RAGE, glycated dendriplexes acted as efficient transfection reagents in CHO-k1 cells which stably express recombinant RAGE. In addition, preincubation with BSA-AGEs, a natural ligand of the RAGE, or dansyl cadaverine, an inhibitor of the RAGE internalization, blocked transfection, confirming their specificity toward RAGE. The results were confirmed in NRK and RAW264.7 cell lines, which naturally express the receptor. The glycated compounds retain their transfection efficiency in the presence of serum and promote in vivo transfection in a mouse model. Accordingly, RAGE is a suitable molecular target for the development of site-directed engineered glycated nonviral gene vectors.
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Dendrímeros/química , Técnicas de Transferência de Genes , Engenharia Genética , Vetores Genéticos/química , Polímeros/química , Receptores Imunológicos/química , Animais , Células CHO , Cadaverina/análogos & derivados , Cadaverina/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cricetulus , Dendrímeros/administração & dosagem , Dendrímeros/síntese química , Dendrímeros/farmacologia , Relação Dose-Resposta a Droga , Feminino , Vetores Genéticos/síntese química , Vetores Genéticos/farmacologia , Camundongos , Modelos Biológicos , Tamanho da Partícula , Polímeros/síntese química , Polímeros/farmacologia , Ratos , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/antagonistas & inibidores , Receptores Imunológicos/metabolismo , Relação Estrutura-Atividade , Propriedades de SuperfícieRESUMO
Cyclodextrins have been conjugated to target various receptors and have also been functionalized with carbohydrates for targeting specific organs. However, this approach is based on a rigid design that implies the ad hoc synthesis of each cyclodextrin-targeting agent conjugate. We hypothesized that: 1)a modular design that decouples the carrier function from the targeting function leads to a flexible system, 2)â combining the reactivity of the vinyl sulfone group toward biomolecules that act as targeting agents with the ability of cyclodextrin to form complexes with a wide range of drugs may yield a versatile system that allows the targeting of different organs with different drugs, and 3) the higher reactivity of histidine residues toward the vinyl sulfone group can be exploited to couple the cyclodextrin to the targeting system with a degree of regioselectivity. As a proof of concept, we synthesized a monovinyl sulfone ß-cyclodextrin (module responsible for the payload), which, after coupling to recombinant antibody fragments raised against Trypanosoma brucei (module responsible for targeting) and loading with nitrofurazone (module responsible for therapeutic action) resulted in an effective delivery system that targets the surface of the parasites and shows trypanocidal activity.
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Portadores de Fármacos/química , Nitrofurazona/administração & dosagem , Sulfonas/química , Tripanossomicidas/administração & dosagem , beta-Ciclodextrinas/química , Anticorpos Imobilizados/química , Anticorpos Imobilizados/imunologia , Sistemas de Liberação de Medicamentos , Humanos , Nitrofurazona/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/efeitos dos fármacos , Trypanosoma brucei brucei/imunologia , Tripanossomíase Africana/tratamento farmacológicoRESUMO
BACKGROUND: S-nitrosylaton is implicated in the regulation of numerous signaling pathways with a diversity of regulatory roles. The high lability of the S-NO bond makes the study of proteins regulated by S-nitrosylation/denitrosylation a challenging task and most studies have focused on already S-nitrosylated proteins. We hypothesize that: i) S-nitrosoglutathione (GSNO) transnitrosylation is a feasible mechanism to account for the physiological S-nitrosylation of rather electropositive sulfur atoms from proteins, ii) affinity chromatography is a suitable approach to isolate proteins that are prone to undergo S-transnitrosylation and iii) vinyl sulfone silica is a suitable chromatographic bead. RESULTS: The combination of vinyl sulfone silica with GSNO yielded an affinity resin that withstood high ionic strength without shrinking or deforming and that it was suitable to isolate potential GSNO transnitrosylation target candidates. Fractions eluted at 1500 mM NaCl resulted in a symmetrical peak for both, protein and S-nitrosothiols, supporting the idea of transnitrosylation by GSNO as a selective process that involves strong and specific interactions with the target protein. Proteomic analysis led to the identification of 22 physiological significant enzymes that differ with the tissue analyzed, being regulatory proteins the most abundant group in hypocotyls. The identification of chloroplastidic FBPase, proteasome, GTP-binding protein, heat shock Hsp70, syntaxin, catalase I, thioredoxin peroxidase and cytochrome P450 that have already been reported as S-nitrosylated by other techniques can be considered as internal positive controls that validate our experimental approach. An additional validation was provided by the prediction of the S-nitrosylation sites in 19 of the GSNO transnitrosylation target candidates. CONCLUSIONS: Vinyl sulfone silica is an open immobilization support that can be turned ad hoc and in a straightforward manner into an affinity resin. Its potential in omic sciences was successfully put to test in the context of the analysis of post-translational modification by S-nitrosylation with two different tissues: mature pea leaves and embryogenic sunflower hypocotyls. The identified proteins reveal an intriguing overlap among S-nitrosylation and both tyrosine nitration and thioredoxin regulation. Chloroplastidic FBPase is a paradigm of such overlap of post-translational modifications since it is reversible modified by thioredoxin and S-nitrosylation and irreversibly by tyrosine nitration. Our results suggest a complex interrelation among different modulation mechanisms mediated by NO-derived molecules.
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Cromatografia de Afinidade/métodos , Helianthus/metabolismo , Pisum sativum/metabolismo , Proteínas de Plantas/metabolismo , S-Nitrosoglutationa/metabolismo , Motivos de Aminoácidos , Cromatografia de Afinidade/instrumentação , Helianthus/química , Pisum sativum/química , Proteínas de Plantas/química , Processamento de Proteína Pós-Traducional , Dióxido de Silício/química , Sulfonas/químicaRESUMO
A new approach for the preparation of carbohydrate-coated magnetic nanoparticles is reported. In a first step, we show that the pH-driven assembly-disassembly natural process that occurs in apoferritin protein is effective for the encapsulation of maghemite nanoparticles of different sizes: 4 and 6 nm. In a second step, we demonstrate that the presence of functional amine groups in the outer shell of apoferritin allows functionalization with two carbohydrates, N-acetyl-D-glucosamine and d-mannose. High-resolution electron microscopy (HREM), high angle annular dark field scanning electron microscopy (HAADF-STEM), electron energy loss spectroscopy (EELS), X-ray diffraction (XRD), and SQUID technique have been used to characterize the magnetic samples, termed herein Apomaghemites. The in vivo magnetic resonance imaging (MRI) studies showed the efficiency in contrasting images for these samples; that is, the r(2) NMR relaxivities are comparable with Endorem (a commercial superparamagnetic MRI contrast agent). The r(2) relaxivity values as well as the pre-contrast and post-contrast T(2)*-weighted images suggested that our systems could be used as perspective superparamagnetic contrast agents for magnetic resonance imaging (MRI). The carbohydrate-functionalized Apomaghemite nanoparticles retained their recognition abilities, as demonstrated by the strong affinity with their corresponding carbohydrate-binding lectins.
Assuntos
Apoferritinas/química , Compostos Férricos/química , Lectinas/química , Magnetismo , Nanopartículas/química , Subunidades Proteicas/química , Carboidratos/química , Cristalografia por Raios X , Concentração de Íons de Hidrogênio , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Icosahedral macromolecules have a wide spectrum of potential nanotechnological applications, the success of which relies on the level of accuracy at which the molecular structure is known. Lumazine synthase from Bacillus subtilis forms a 150 A icosahedral capsid consisting of 60 subunits and crystallizes in space group P6(3)22 or C2. However, the quality of these crystals is poor and structural information is only available at 2.4 A resolution. As classical strategies for growing better diffracting crystals have so far failed, protein engineering has been employed in order to improve the overexpression and purification of the molecule as well as to obtain new crystal forms. Two cysteines were replaced to bypass misfolding problems and a charged surface residue was replaced to force different molecular packings. The mutant protein crystallizes in space group R3, with unit-cell parameters a = b = 313.02, c = 365.77 A, alpha = beta = 90.0, gamma = 120 degrees , and diffracts to 1.6 A resolution.
Assuntos
Complexos Multienzimáticos/normas , Engenharia de Proteínas/normas , Bacillus subtilis/enzimologia , Bacillus subtilis/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/normas , Cristalização/métodos , Cristalização/normas , Complexos Multienzimáticos/química , Complexos Multienzimáticos/genética , Mutagênese Sítio-Dirigida/métodos , Mutagênese Sítio-Dirigida/normas , Engenharia de Proteínas/métodos , Riboflavina Sintase/química , Riboflavina Sintase/genética , Riboflavina Sintase/normasRESUMO
The crystallization of glycoproteins is one of the challenges to be confronted by the crystallographic community in the frame of what is known as glycobiology. The state of the art for the crystallization of glycoproteins is not promising and removal of the carbohydrate chains is generally suggested since they are flexible and a source of heterogeneity. In this paper, the feasibility of introducing glucose into the model protein hen egg-white lysozyme via a post-purification glycosylation reaction that may turn any protein into a model glycoprotein whose carbohydrate fraction can be manipulated is demonstrated.
Assuntos
Glicosilação , Muramidase/química , Animais , Precipitação Química , Galinhas , Cristalização/métodos , Proteínas do Ovo , Difração de Raios XRESUMO
Concanavalin A has been crystallized in the presence of the ligand (6-S-beta-D-galactopyranosyl-6-thio)-cyclomaltoheptaose. The crystals are isomorphous to those reported for ConA complexed with peptides at low resolution (3.00-2.75 angstroms). The structure was solved at 1.9 angstroms, with free R and R values of 0.201 and 0.184, respectively. As expected, no molecules of the ligand were bound to the protein. Soaking in the cryobuffer left its fingerprint as 25 molecules of glycerol in the bound solvent, most of them at specific positions. The fact that a glycerol molecule is located in the sugar-binding pocket of each of the four subunits in the asymmetric unit and another is located in two of the peptide-binding sites suggests a recognition phenomenon rather than a displacement of water molecules by glycerol. Crystal contact analysis shows that a relation exists between the residues that form hydrogen bonds to other asymmetric units and the space group: contact Asp58-Ser62 is a universal feature of ConA crystals, while Ser66-His121, Asn69-Asn118 and Tyr100-His205 contacts are general features of the C222(1) crystal form.
Assuntos
Concanavalina A/química , Cristalografia por Raios X/métodos , Ciclodextrinas/química , Azidas/química , Sítios de Ligação , Carboidratos/química , Portadores de Fármacos , Glicerol/química , Ligação de Hidrogênio , Concentração de Íons de Hidrogênio , Íons , Manganês/química , Modelos Moleculares , Conformação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de ProteínaRESUMO
Desde el descubrimiento de la penicilina, hace ya casi tres cuartos de siglo, se han venido produciendo numerosos antibióticos para el tratamiento de enfermedades infecciosas. Esto ha dado como resultado una importante disminución de este tipo de enfermedades y, en consecuencia, del número de muertes, sin embargo esta nueva generación de fármacos trajo como resultado un uso generalizado e indiscriminado que promovió el desarrollo de la resistencia bacteriana. Actualmente muchas investigaciones en el mundo buscan nuevas y efectivas propuestas para el manejo de las infecciones baterianas y desarrollan interesantes alternativas de solución. El secuenciamiento del genoma bacteriano para conocer blancos moleculares, el uso de bacteriófagos y la búsqueda de productos de origen natural y sintético que representan una fuente rica en compuestos con actividad biológica, son sólo una pequeña muestra de los diversos campos que se pueden explorar para encontrar una solución al problema de la resistencia bacteriana