RESUMO
The dihydroneopterin aldolase (DHNA, EC 4.1.2.25) activity of FolB protein is required for the conversion of 7,8-dihydroneopterin (DHNP) to 6-hydroxymethyl-7,8-dihydropterin (HP) and glycolaldehyde (GA) in the folate pathway. FolB protein from Mycobacterium tuberculosis (MtFolB) is essential for bacilli survival and represents an important molecular target for drug development. S8-functionalized 8-mercaptoguanine derivatives were synthesised and evaluated for inhibitory activity against MtFolB. The compounds showed IC50 values in the submicromolar range. The inhibition mode and inhibition constants were determined for compounds that exhibited the strongest inhibition. Additionally, molecular docking analyses were performed to suggest enzyme-inhibitor interactions and ligand conformations. To the best of our knowledge, this study describes the first class of MtFolB inhibitors.
Assuntos
Aldeído Liases/antagonistas & inibidores , Antibacterianos/farmacologia , Inibidores Enzimáticos/farmacologia , Guanosina/análogos & derivados , Simulação de Acoplamento Molecular , Mycobacterium tuberculosis/efeitos dos fármacos , Tionucleosídeos/farmacologia , Aldeído Liases/genética , Aldeído Liases/metabolismo , Antibacterianos/síntese química , Antibacterianos/química , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Guanosina/síntese química , Guanosina/química , Guanosina/farmacologia , Testes de Sensibilidade Microbiana , Estrutura Molecular , Mycobacterium tuberculosis/enzimologia , Tionucleosídeos/síntese química , Tionucleosídeos/químicaRESUMO
The guanine base in DNA, due to its low oxidation potential, is particularly sensitive to chemical modifications. A large number of guanine lesions have been characterized and studied in some detail due to their relationship with tissue inflammations. Nevertheless, one example of these lesions is the formation of 8-nitro-guanosine, but the NMR data of this compound was only partially interpreted. A comprehensive study of the two possible tautomeric forms, through a detailed characterization of this compound, has implications for its base pairing properties. The target compound was obtained through a synthetic sequence of five steps, where all intermediates were fully characterized using spectral data. The analysis of the two tautomers was then evaluated through NMR spectroscopy and theoretical calculations of the chemical shifts and NH coupling constants, which were also compared with the data from guanosine.
Assuntos
Guanosina/análogos & derivados , Espectroscopia de Ressonância Magnética , Modelos Teóricos , Nitrocompostos/química , DNA/química , Guanosina/síntese química , Guanosina/química , Hidrólise , Espectroscopia de Ressonância Magnética/métodos , Espectrometria de Massas/métodos , Nitrocompostos/síntese químicaRESUMO
The oxidation of the free nucleoside 2'-deoxyguanosine (dGuo) by singlet molecular oxygen ((1)O2) has been studied over the three last decades due to the major role of DNA oxidation products in process such as ageing, mutation and carcinogenesis. In the present work we investigated the dGuo oxidation by (1)O2 in the presence of the important low molecular antioxidant, glutathione, in its reduced (GSH) and oxidized (GSSG) forms. There were applied different conditions of concentration, pH, time of incubation, and the use of a [(18)O]-labeled thermolabile endoperoxide naphthalene derivative as a source of [(18)O]-labeled (1)O2. Data was obtained through high performance liquid chromatography (HPLC) and HPLC coupled to micrOTOF Q-II analysis of the main oxidation products: the diastereomers of spiroiminodihydantoin-2'-deoxyribonucleosides (dSp) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo). An intriguing result was that 8-oxodGuo levels increased by 100 fold when dGuo was oxidized by (1)O2 in the presence of GSH and by 2 fold in the presence of GSSG, while dSp levels dropped to zero for both conditions. All data from dGuo, 8-oxodGuo and dSp quantification together with the analysis of residual GSH/GSSG content in each sample strongly suggest that glutathione modifies the mechanism of dGuo oxidation by (1)O2 by disfavoring the pathway of dSp formation.
Assuntos
Desoxiguanosina/metabolismo , Glutationa/metabolismo , Oxigênio Singlete/metabolismo , 8-Hidroxi-2'-Desoxiguanosina , Desoxiguanosina/análogos & derivados , Desoxiguanosina/química , Glutationa/química , Dissulfeto de Glutationa/química , Dissulfeto de Glutationa/metabolismo , Guanosina/análogos & derivados , Guanosina/química , Guanosina/metabolismo , Técnicas In Vitro , Modelos Químicos , Oxirredução , Oxigênio Singlete/química , Compostos de Espiro/química , Compostos de Espiro/metabolismoRESUMO
OBJECTIVE: To examine the prevalence of blindness, visual impairment, and related eye diseases and conditions among adults in El Salvador, and to explore socioeconomic inequalities in their prevalence by education level and occupational status, stratified by sex. METHODS: Based upon the Rapid Assessment of Avoidable Blindness (RAAB) methodology, this nationwide sample comprised 3 800 participants (3 399 examined) ≥ 50 years old from 76 randomly selected clusters of 50 persons each. The prevalence of blindness, visual impairment and related eye diseases and conditions, including uncorrected refractive error (URE), was calculated for categories of education level and occupational status. Multiple logistic regression models were fitted to calculate odds ratios (ORs) and 95% confidence intervals (CIs) and stratified by sex. RESULTS: Age-adjusted prevalence was 2.4% (95% CI: 2.2-2.6) for blindness (men: 2.8% (95% CI: 2.5-3.1); women: 2.2% (95% CI: 1.9-2.5)) and 11.8% (95% CI: 11.6-12.0) for moderate visual impairment (men: 10.8% (95% CI: 10.5-11.1); women: 12.6% (95% CI: 12.4-12.8)). The proportion of visual impairment due to cataract was 43.8% in men and 33.5% in women. Inverse gradients of socioeconomic inequalities were observed in the prevalence of visual impairment. For example, the age-adjusted OR (AOR) was 3.4 (95% CI: 2.0-6.4) for visual impairment and 4.3 (95% CI: 2.1-10.4) for related URE in illiterate women compared to those with secondary education, and 1.9 (95% CI: 1.1-3.1) in cataract in unemployed men. CONCLUSIONS: Blindness and visual impairment prevalence is high in the El Salvador adult population. The main associated conditions are cataract and URE, two treatable conditions. As socioeconomic and gender inequalities in ocular health may herald discrimination and important barriers to accessing affordable, good-quality, and timely health care services, prioritization of public eye health care and disability policies should be put in place, particularly among women, the unemployed, and uneducated people.
OBJETIVO: Analizar la prevalencia de la ceguera, la deficiencia visual, y las enfermedades y afecciones oculares relacionadas en adultos de El Salvador, y explorar las desigualdades socioeconómicas en cuanto a su prevalencia según el nivel educativo y la situación laboral, estratificados por sexos. MÉTODOS: Se adoptó el método de Evaluación Rápida de la Ceguera Evitable, y se escogió una muestra a escala nacional de 3 800 participantes (de ellos se examinaron 3 399) de 50 años de edad o mayores, pertenecientes a 76 agrupamientos seleccionados aleatoriamente y constituidos por 50 personas cada uno. Se calculó la prevalencia de la ceguera, la deficiencia visual y las enfermedades y afecciones oculares relacionadas, incluido el error de refracción no corregido, según las diferentes categorías de nivel educativo y situación laboral. Se emplearon modelos de regresión logística múltiple para calcular las razones de posibilidades (OR) y los intervalos de confianza (IC) de 95%, y se estratificaron por sexos. RESULTADOS: La prevalencia ajustada por edad fue de 2,4% (IC de 95%: 2,2-2,6) para la ceguera (hombres: 2,8% [IC de 95%: 2,5-3,1]; mujeres: 2,2% [IC de 95%: 1,9-2,5]) y de 11,8% (IC de 95%: 11,6-12,0) para la deficiencia visual moderada (hombres: 10,8% [IC de 95%: 10,5-11,1]; mujeres: 12,6% [IC de 95%: 12,4-12,8]). La proporción de deficiencias visuales debidas a catarata fue de 43,8% en los hombres y de 33,5% en las mujeres. En la prevalencia de la deficiencia visual se observaron gradientes inversos de desigualdades socioeconómicas. Por ejemplo, la OR ajustada por edad fue de 3,4 (IC de 95%: 2,0-6,4) para la deficiencia visual y de 4,3 (IC de 95%: 2,1-10,4) para el error de refracción no corregido relacionado en las mujeres analfabetas, en comparación con las que tenían un nivel de educación secundaria, y fue de 1,9 (IC de 95%: 1,1-3,1) para la catarata en los hombres desempleados. CONCLUSIONES: La prevalencia de ceguera y deficiencia visual es alta en la población adulta de El Salvador. Las principales afecciones asociadas son la catarata y el error de refracción no corregido, ambas tratables. Puesto que las desigualdades socioeconómicas y de género en materia de salud ocular pueden ser indicativas de discriminación y de la existencia de barreras importantes para obtener acceso a servicios de atención de salud asequibles, de buena calidad y oportunos, es preciso dar prioridad a la atención oftalmológica pública y a las políticas dirigidas a corregir la discapacidad, en particular en las mujeres y en las personas desempleadas y sin formación.
Assuntos
Carcinógenos/química , Carcinógenos/síntese química , Adutos de DNA/biossíntese , Adutos de DNA/química , Compostos de Epóxi/química , Compostos de Epóxi/síntese química , Guanosina/química , Adutos de DNA/efeitos dos fármacos , Estabilidade de Medicamentos , Compostos de Epóxi/toxicidade , Cinética , Espectrometria de Massas , EstereoisomerismoRESUMO
Oligonucleotides have unique molecular recognition properties, being involved in biological mechanisms such as cell-surface receptor recognition or gene silencing. For their use in human therapy for drug or gene delivery, the cell membrane remains a barrier, but this can be obviated by grafting a hydrophobic tail to the oligonucleotide. Here we demonstrate that two oligonucleotides, one consisting of 12 guanosine units (G(12)), and the other one consisting of five adenosine and seven guanosine (A(5)G(7)) units, when functionalized with poly(butadiene), namely PB-G(12) and PB-A(5)G(7), can be inserted into Langmuir monolayers of dipalmitoyl phosphatidyl choline (DPPC), which served as a cell membrane model. PB-G(12) and PB-A(5)G(7) were found to affect the DPPC monolayer even at high surface pressures. The effects from PB-G(12) were consistently stronger, particularly in reducing the elasticity of the DPPC monolayers, which may have important biological implications. Multilayers of DPPC and nucleotide-based copolymers could be adsorbed onto solid supports, in the form of Y-type LB films, in which the molecular-level interaction led to lower energies in the vibrational spectra of the nucleotide-based copolymers. This successful deposition of solid films opens the way for devices to be produced which exploit the molecular recognition properties of the nucleotides.
Assuntos
Membrana Celular/química , Membrana Celular/metabolismo , Modelos Biológicos , Oligonucleotídeos/química , Oligonucleotídeos/metabolismo , 1,2-Dipalmitoilfosfatidilcolina/química , 1,2-Dipalmitoilfosfatidilcolina/metabolismo , Adenosina/química , Adenosina/metabolismo , Butadienos/química , Elasticidade , Elastômeros/química , Guanosina/química , Guanosina/metabolismo , Espectrofotometria Ultravioleta , Espectroscopia de Infravermelho com Transformada de Fourier , Tensão Superficial , Fatores de TempoRESUMO
Here we show 2'-deoxyguanosine derivatives that self-assemble in aqueous media into discrete supramolecular hexadecamers and exhibit the lower critical solution temperature (LCST) phenomenon. Spectroscopic, calorimetric, and electron microscopy studies support the fact that above the transition temperature (T(t)) the supramolecules further assemble into nanoscopic spherical globules of low polydispersity. Furthermore, the T(t) can be tuned to higher values by the addition of a more hydrophilic derivative. These findings uncover a new paradigm in the development of smart thermosensitive materials with properties and applications complementary to those of polymers.
Assuntos
Desoxiguanosina/análogos & derivados , Guanosina/análogos & derivados , Temperatura Alta , Guanosina/química , Polímeros/química , Água/químicaRESUMO
Cellular nucleic acid binding protein (CNBP) is a small single-stranded nucleic acid binding protein made of seven Zn knuckles and an Arg-Gly rich box. CNBP is strikingly conserved among vertebrates and was reported to play broad-spectrum functions in eukaryotic cells biology. Neither its biological function nor its mechanisms of action were elucidated yet. The main goal of this work was to gain further insights into the CNBP biochemical and molecular features. We studied Bufo arenarum CNBP (bCNBP) binding to single-stranded nucleic acid probes representing the main reported CNBP putative targets. We report that, although bCNBP is able to bind RNA and single-stranded DNA (ssDNA) probes in vitro, it binds RNA as a preformed dimer whereas both monomer and dimer are able to bind to ssDNA. A systematic analysis of variant probes shows that the preferred bCNBP targets contain unpaired guanosine-rich stretches. These data expand the knowledge about CNBP binding stoichiometry and begins to dissect the main features of CNBP nucleic acid targets. Besides, we show that bCNBP presents a highly disordered predicted structure and promotes the annealing and melting of nucleic acids in vitro. These features are typical of proteins that function as nucleic acid chaperones. Based on these data, we propose that CNBP may function as a nucleic acid chaperone through binding, remodeling, and stabilizing nucleic acids secondary structures. This novel CNBP biochemical activity broadens the field of study about its biological function and may be the basis to understand the diverse ways in which CNBP controls gene expression.
Assuntos
DNA de Cadeia Simples/metabolismo , Proteínas de Ligação a DNA/metabolismo , Quadruplex G , Chaperonas Moleculares/metabolismo , Proteínas de Ligação a RNA/metabolismo , Regiões 5' não Traduzidas/química , Regiões 5' não Traduzidas/metabolismo , Animais , Sequência de Bases , Bufo arenarum , Sondas de DNA/química , Sondas de DNA/metabolismo , DNA de Cadeia Simples/química , Proteínas de Ligação a DNA/química , Embrião não Mamífero/química , Embrião não Mamífero/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Guanosina/química , Guanosina/metabolismo , Humanos , Dados de Sequência Molecular , Ligação Proteica , Biossíntese de Proteínas , Sondas RNA/química , Sondas RNA/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Xenopus laevis/metabolismo , Dedos de ZincoRESUMO
The main singlet molecular oxygen ((1)O(2)) oxidation products of free 2'-deoxyguanosine (dGuo) in aqueous solution were identified as a pair of diastereomeric spiroiminodihydantoin 2'-deoxyribonucleosides (dSp) together with 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo). In the present work, evidence is provided from (18)[(1)O(2)] and H(2) (18)O labeling experiments, using HPLC-ESI-MS/MS, that the formation of dSp is explained by the addition of water to a reactive quinonoid intermediate, and a second reaction pathway leading to dSp involves (1)O(2) oxidation of initially generated 8-oxodGuo.
Assuntos
Desoxiguanosina/química , Guanosina/análogos & derivados , Compostos de Espiro/química , Cromatografia Líquida de Alta Pressão , Guanosina/química , Hidantoínas/química , Cinética , Estrutura Molecular , Oxirredução , Isótopos de Oxigênio , Oxigênio Singlete , Soluções , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem , ÁguaRESUMO
Transducin (T) mediates vision in retinal rods by transmitting light signals detected by rhodopsin to a cGMP phosphodiesterase. The flow of information relies on a subunit association/dissociation cycle of T regulated by a guanine nucleotide exchange/hydrolysis reaction. 5'-[p-(Fluorosulfonyl)benzoyl] guanosine (FSBG) was synthesized and examined here as an affinity label for the guanine nucleotide binding site of T. Although the relative binding affinity of FSBG to T was much lower than for GTP and beta,gamma-imido-guanosine 5'-triphosphate (GMPPNP), the incorporation of FSBG to T inhibited its light-dependent [(3)H] GMPPNP binding activity in a concentration dependent manner. Additionally, GDP, GTP and GTP analogs hindered the binding of [(3)H] FSBG to T. These results demonstrated that FSBG could be used to specifically modify the active site of T. In addition, FSBG was not capable of dissociating T from T:photoactivated rhodopsin complexes, suggesting that in this case FSBG is acting as a GDP analog.
Assuntos
Marcadores de Afinidade , Guanina/metabolismo , Guanosina/análogos & derivados , Transducina/química , Transducina/metabolismo , Animais , Sítios de Ligação , Bovinos , Guanina/química , Guanosina/química , Guanosina/metabolismo , Ligação ProteicaRESUMO
Purine nucleoside phosphorylase (PNP) is a key enzyme in the purine-salvage pathway, which allows cells to utilize preformed bases and nucleosides in order to synthesize nucleotides. PNP is specific for purine nucleosides in the beta-configuration and exhibits a strong preference for purines containing a 6-keto group and ribosyl-containing nucleosides relative to the corresponding analogues. PNP was crystallized in complex with ligands and data collection was performed using synchrotron radiation. This work reports the structure of human PNP in complex with guanosine (at 2.80 A resolution), 3'-deoxyguanosine (at 2.86 A resolution) and 8-azaguanine (at 2.85 A resolution). These structures were compared with the PNP-guanine, PNP-inosine and PNP-immucillin-H complexes solved previously.