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1.
PLoS Genet ; 16(10): e1009085, 2020 10.
Artículo en Inglés | MEDLINE | ID: mdl-33125364

RESUMEN

DNA supercoiling is essential for all living cells because it controls all processes involving DNA. In bacteria, global DNA supercoiling results from the opposing activities of topoisomerase I, which relaxes DNA, and DNA gyrase, which compacts DNA. These enzymes are widely conserved, sharing >91% amino acid identity between the closely related species Escherichia coli and Salmonella enterica serovar Typhimurium. Why, then, do E. coli and Salmonella exhibit different DNA supercoiling when experiencing the same conditions? We now report that this surprising difference reflects disparate activation of their DNA gyrases by the polyamine spermidine and its precursor putrescine. In vitro, Salmonella DNA gyrase activity was sensitive to changes in putrescine concentration within the physiological range, whereas activity of the E. coli enzyme was not. In vivo, putrescine activated the Salmonella DNA gyrase and spermidine the E. coli enzyme. High extracellular Mg2+ decreased DNA supercoiling exclusively in Salmonella by reducing the putrescine concentration. Our results establish the basis for the differences in global DNA supercoiling between E. coli and Salmonella, define a signal transduction pathway regulating DNA supercoiling, and identify potential targets for antibacterial agents.


Asunto(s)
Girasa de ADN/genética , ADN-Topoisomerasas de Tipo I/genética , ADN Superhelicoidal/genética , Escherichia coli/genética , Salmonella typhimurium/genética , Girasa de ADN/efectos de los fármacos , ADN-Topoisomerasas de Tipo I/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Escherichia coli/enzimología , Magnesio/farmacología , Putrescina/farmacología , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/enzimología , Espermidina/biosíntesis
2.
Inorg Chem ; 59(20): 14866-14870, 2020 Oct 19.
Artículo en Inglés | MEDLINE | ID: mdl-32993282

RESUMEN

Antimicrobial photodynamic therapy (APDT) has gained increased attention because of its broad spectrum activity and lower likelihood to elicit bacterial resistance. Although many photosensitizers excel at eradicating Gram-positive bacterial infections, they are generally less potent when utilized against Gram-negative bacteria. We hypothesized that conjugating the DNA-targeting, antimicrobial peptide buforin II to a metal-based photosensitizer would result in a potent APDT agent. Herein, we present the synthesis and characterization of a buforin II-[Ru(bpy)3]2+ bioconjugate (1). The submicromolar activity of 1 against the multidrug-resistant strains Escherichia coli AR 0114 and Acinetobacter baumannii Naval-17 indicates strong synergy between the ruthenium complex and buforin II. Our mechanistic studies point to an increased rate of DNA damage by 1 compared to [Ru(bpy)3]2+. These results suggest that conjugating metal complexes to antimicrobial peptides can lead to potent antimicrobial agents.


Asunto(s)
Antibacterianos/farmacología , Complejos de Coordinación/farmacología , Farmacorresistencia Bacteriana/efectos de los fármacos , Resistencia a Múltiples Medicamentos/efectos de los fármacos , Fármacos Fotosensibilizantes/farmacología , Proteínas/farmacología , Antibacterianos/síntesis química , Antibacterianos/efectos de la radiación , Complejos de Coordinación/síntesis química , Complejos de Coordinación/efectos de la radiación , Daño del ADN/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Bacterias Gramnegativas/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Fármacos Fotosensibilizantes/síntesis química , Fármacos Fotosensibilizantes/efectos de la radiación , Proteínas/síntesis química , Rutenio/química , Rutenio/efectos de la radiación , Oxígeno Singlete/metabolismo
3.
Sci Rep ; 9(1): 18216, 2019 12 03.
Artículo en Inglés | MEDLINE | ID: mdl-31796849

RESUMEN

The World Health Organization has listed C. jejuni as one of 12 microorganisms on a global priority list for antibiotic resistance due to a rapid increase in strains resistant to fluoroquinolone antibiotics. This fluoroquinolone resistance is conferred through a single point mutation in the QRDR region within the gyrA gene known to be involved in DNA supercoiling. We have previously revealed that changes in DNA supercoilikng play a major role in the regulation of virulence in C. jejuni with relaxation of DNA supercoiling associated with increased attachment to and invasion of human epithelial cells. The aim of this study was to investigate whether fluoroquinolone resistant strains of C. jejuni displayed altered supercoiling associated phenotypes. A panel of fluoroquinolone resistant mutants were derived and shown to have a greater ability to form viable biofilms under aerobic conditions, invade epithelial cells and promote virulence in the Galleria mellonella model of infection. We thus report for the first time that fluoroquinolone resistance in C. jejuni is associated with an increase in virulence and the ability to form viable biofilms in oxygen rich environments. These altered phenotypes likely play a critical role in the continued increase in fluoroquinolone resistance observed for this important pathogen.


Asunto(s)
Biopelículas/crecimiento & desarrollo , Infecciones por Campylobacter/tratamiento farmacológico , Campylobacter jejuni/patogenicidad , Farmacorresistencia Bacteriana/genética , Fluoroquinolonas/farmacología , Infecciones por Campylobacter/microbiología , Campylobacter jejuni/genética , Girasa de ADN/genética , Girasa de ADN/metabolismo , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/metabolismo , Farmacorresistencia Bacteriana/efectos de los fármacos , Fluoroquinolonas/uso terapéutico , Células HT29 , Humanos , Pruebas de Sensibilidad Microbiana , Mutación Puntual/efectos de los fármacos , Virulencia/genética
4.
Sci Rep ; 9(1): 4486, 2019 03 14.
Artículo en Inglés | MEDLINE | ID: mdl-30872616

RESUMEN

Temperature shifts trigger genome-wide changes in Escherichia coli's gene expression. We studied if chromosome integration impacts on a gene's sensitivity to these shifts, by comparing the single-RNA production kinetics of a PLacO3O1 promoter, when chromosomally-integrated and when single-copy plasmid-borne. At suboptimal temperatures their induction range, fold change, and response to decreasing temperatures are similar. At critically low temperatures, the chromosome-integrated promoter becomes weaker and noisier. Dissection of its initiation kinetics reveals longer lasting states preceding open complex formation, suggesting enhanced supercoiling buildup. Measurements with Gyrase and Topoisomerase I inhibitors suggest hindrance to escape supercoiling buildup at low temperatures. Consistently, similar phenomena occur in energy-depleted cells by DNP at 30 °C. Transient, critically-low temperatures have no long-term consequences, as raising temperature quickly restores transcription rates. We conclude that the chromosomally-integrated PLacO3O1 has higher sensitivity to low temperatures, due to longer-lasting super-coiled states. A lesser active, chromosome-integrated native lac is shown to be insensitive to Gyrase overexpression, even at critically low temperatures, indicating that the rate of escaping positive supercoiling buildup is temperature and transcription rate dependent. A genome-wide analysis supports this, since cold-shock genes exhibit atypical supercoiling-sensitivities. This phenomenon might partially explain the temperature-sensitivity of some transcriptional programs of E. coli.


Asunto(s)
Cromosomas Bacterianos/genética , Escherichia coli/crecimiento & desarrollo , Perfilación de la Expresión Génica/métodos , Plásmidos/genética , Frío , ADN Superhelicoidal/efectos de los fármacos , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Regiones Promotoras Genéticas , Imagen Individual de Molécula , Estrés Fisiológico , Inhibidores de Topoisomerasa I/farmacología , Inhibidores de Topoisomerasa II/farmacología , Secuenciación Completa del Genoma
5.
J Photochem Photobiol B ; 177: 112-123, 2017 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-29089229

RESUMEN

As a promising next-generation photodynamic therapy (PDT) photosensitizer, TiO2 nanoparticles (NPs) has gained great attention due to its higher efficiency. Yet, its application in PDT is strongly limited by its UV light response range. In this work, TiO2 NPs conjugated with reduced graphene oxide (RGO-TiO2) composites were successfully prepared by hydrothermal reduction method. They were characterized by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), Transmission electron microscope (TEM), Brunauer-Emmett-Teller (BET), UV-vis spectroscopy and X-ray photoelectron spectroscopy (XPS). Superior adsorption and killing efficiency under UV-A light or visible light were achieved in the presence of the RGO rather than that of unmodified TiO2. The optimal photocatalytic activity was obtained when modified proportion was 0.2 (RGO:TiO2). Dark cytotoxicity was observed using 0-500µgmL-1 RGO-TiO2 during long incubation time. In parallel, following exposure of human hepatocellular carcinoma cell line (HepG2 cells) to RGO-TiO2 and UV-A or visible light irradiation, a marked decrease in the ratio of the super-coiled DNA, mitochondrial membrane potential (MMP), and the oxidative damage effects, as well as increased the apoptosis rate and intracellular calcium concentration were observed. Moreover, photocatalytic RGO-TiO2 composites killed the HepG2 cells by apoptosis pathway. The results suggested that RGO-TiO2 composites were an excellent candidate as a PDT photosensitizer in the near future.


Asunto(s)
Grafito/química , Nanocompuestos/química , Óxidos/farmacología , Titanio/química , Titanio/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Calcio/metabolismo , Catálisis , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/efectos de la radiación , Grafito/farmacología , Células Hep G2 , Humanos , Luz , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de la radiación , Nanopartículas del Metal/química , Microscopía Electrónica de Transmisión , Nanocompuestos/toxicidad , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/efectos de la radiación , Óxidos/química , Espectroscopía de Fotoelectrones , Espectroscopía Infrarroja por Transformada de Fourier , Rayos Ultravioleta , Difracción de Rayos X
6.
Molecules ; 22(10)2017 Sep 30.
Artículo en Inglés | MEDLINE | ID: mdl-28973991

RESUMEN

Phenolics in food and fruit tree leaves exist in free, soluble-conjugate, and insoluble-bound forms. In this study, in order to enhance the bioavailability of insoluble-bound phenolics from guava leaves (GL), the ability of enzyme-assisted extraction in improving the release of insoluble-bound phenolics was investigated. Compared to untreated GL, single xylanase-assisted extraction did not change the composition and yield of soluble phenolics, whereas single cellulase or ß-glucosidase-assisted extraction significantly enhanced the soluble phenolics content of PGL. However, complex enzyme-assisted extraction (CEAE) greatly improved the soluble phenolics content, flavonoids content, ABTS, DPPH, and FRAP by 103.2%, 81.6%, 104.4%, 126.5%, and 90.3%, respectively. Interestingly, after CEAE, a major proportion of phenolics existed in the soluble form, and rarely in the insoluble-bound form. Especially, the contents of quercetin and kaempferol with higher bio-activity were enhanced by 3.5- and 2.2-fold, respectively. More importantly, total soluble phenolics extracts of GL following CEAE exhibited the highest antioxidant activity and protective effect against supercoiled DNA damage. This enzyme-assisted extraction technology can be useful for extracting biochemical components from plant matrix, and has good potential for use in the food and pharmaceutical industries.


Asunto(s)
Antioxidantes/aislamiento & purificación , Complejos Multienzimáticos/metabolismo , Fenoles/aislamiento & purificación , Extractos Vegetales/química , Hojas de la Planta/química , Antioxidantes/farmacología , Cromatografía Líquida de Alta Presión/métodos , Daño del ADN/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Flavonoides/aislamiento & purificación , Flavonoides/farmacología , Frutas/química , Humanos , Psidium/química , Solubilidad , Agua/química
7.
Nucleic Acids Res ; 44(15): 7292-303, 2016 09 06.
Artículo en Inglés | MEDLINE | ID: mdl-27378778

RESUMEN

We studied the transcriptional response to an increase in DNA supercoiling in Streptococcus pneumoniae by using seconeolitsine, a new topoisomerase I inhibitor. A homeostatic response allowing recovery of supercoiling was observed in cells treated with subinhibitory seconeolitsine concentrations. Supercoiling increases of 40.7% (6 µM) and 72.9% (8 µM) were lowered to 8.5% and 44.1%, respectively. Likewise, drug removal facilitated the recovery of cell viability and DNA-supercoiling. Transcription of topoisomerase I depended on the supercoiling level. Also specific binding of topoisomerase I to the gyrase A gene promoter was detected by chromatin-immunoprecipitation. The transcriptomic response to 8 µM seconeolitsine had two stages. An early stage, associated to an increase in supercoiling, affected 10% of the genome. A late stage, manifested by supercoiling recovery, affected 2% of the genome. Nearly 25% of the early responsive genes formed 12 clusters with a coordinated transcription. Clusters were 6.7-31.4 kb in length and included 9-22 responsive genes. These clusters partially overlapped with those observed under DNA relaxation, suggesting that bacteria manage supercoiling stress using pathways with common components. This is the first report of a coordinated global transcriptomic response that is triggered by an increase in DNA supercoiling in bacteria.


Asunto(s)
ADN-Topoisomerasas de Tipo I/genética , ADN Superhelicoidal/genética , Homeostasis/genética , Familia de Multigenes , Streptococcus pneumoniae/genética , Benzodioxoles/farmacología , Girasa de ADN/genética , ADN-Topoisomerasas de Tipo I/biosíntesis , ADN-Topoisomerasas de Tipo I/metabolismo , ADN Bacteriano/efectos de los fármacos , ADN Bacteriano/genética , ADN Superhelicoidal/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Genes Bacterianos/genética , Homeostasis/efectos de los fármacos , Viabilidad Microbiana/efectos de los fármacos , Viabilidad Microbiana/genética , Fenantrenos/farmacología , Streptococcus pneumoniae/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Transcriptoma/efectos de los fármacos , Transcriptoma/genética
8.
Bioorg Med Chem Lett ; 26(2): 322-327, 2016 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-26711148

RESUMEN

Three platinum(II) complexes of (1R,2R)-N(1)-cyclopentyl-1,2-cyclohexanediamine with malonate derivatives were designed, synthesized and spectrally characterized. MTT assay showed that the complexes possessed positive cytotoxic effect on the four human solid tumor cell lines. Among the complexes, complex 2 demonstrated the strongest cytotoxic activity compared to cisplatin and oxaliplatin against HepG2 cell line (IC50=3.04µM). Furthermore, the results of gel electrophoresis revealed that complex 2 interacted with DNA in a different mode from that of cisplatin. Mechanism studies of cell proliferation inhibition and cellular uptake indicated that complex 2 entered HepG2 cell more efficiently than cisplatin, exhibited massive G2 accumulation and then induced apoptosis.


Asunto(s)
Antineoplásicos/farmacología , Complejos de Coordinación/farmacología , Ciclohexilaminas/farmacología , Malonatos/farmacología , Compuestos Organoplatinos/farmacología , Platino (Metal)/química , Antineoplásicos/síntesis química , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Complejos de Coordinación/síntesis química , Complejos de Coordinación/química , Ciclohexilaminas/síntesis química , Ciclohexilaminas/química , División del ADN/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ligandos , Malonatos/síntesis química , Malonatos/química , Compuestos Organoplatinos/síntesis química , Relación Estructura-Actividad
9.
Biomed Res Int ; 2015: 320490, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26682217

RESUMEN

The effects of dimethyl sulfoxide (DMSO) on supercoiled plasmid DNA relaxation catalyzed by two typical type I topoisomerases were investigated in our studies. It is shown that DMSO in a low concentration (less than 20%, v/v) can induce a dose-related enhancement of the relaxation efficiency of Escherichia coli topoisomerase I (type IA). Conversely, obvious inhibitory effect on the activity of calf thymus topoisomerase I (type IB) was observed when the same concentration of DMSO is used. In addition, our studies demonstrate that 20% DMSO has an ability to reduce the inhibitory effect on EcTopo I, which was induced by double-stranded oligodeoxyribonucleotides while the same effect cannot be found in the case of CtTopo I. Moreover, our AFM examinations suggested that DMSO can change the conformation of negatively supercoiled plasmid by creating some locally loose regions in DNA molecules. Combining all the lines of evidence, we proposed that DMSO enhanced EcTopo I relaxation activity by (1) increasing the single-stranded DNA regions for the activities of EcTopo I in the early and middle stages of the reaction and (2) preventing the formation of double-stranded DNA-enzyme complex in the later stage, which can elevate the effective concentration of the topoisomerase in the reaction solution.


Asunto(s)
Catálisis/efectos de los fármacos , ADN-Topoisomerasas de Tipo I/metabolismo , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/metabolismo , ADN/efectos de los fármacos , Dimetilsulfóxido/farmacología , ADN/metabolismo , ADN de Cadena Simple/efectos de los fármacos , ADN de Cadena Simple/metabolismo , Escherichia coli/metabolismo , Plásmidos/metabolismo
10.
Oncotarget ; 6(11): 8960-73, 2015 Apr 20.
Artículo en Inglés | MEDLINE | ID: mdl-25840421

RESUMEN

Both microtubule and topoisomerase II (Top2) are important anticancer targets and their respective inhibitors are widely used in combination for cancer therapy. However, some combinations could be mutually antagonistic and drug resistance further limits their therapeutic efficacy. Here we report YCH337, a novel α-carboline derivative that targets both microtubule and Top2, eliciting tumor proliferation and growth inhibition and overcoming drug resistance. YCH337 inhibited microtubule polymerization by binding to the colchicine site and subsequently led to mitotic arrest. It also suppressed Top2 and caused DNA double-strand breaks. It disrupted microtubule more potently than Top2. YCH337 induced reversible mitotic arrest at low concentrations but persistent DNA damage. YCH337 caused intrinsic and extrinsic apoptosis and decreased MCL-1, cIAP1 and XIAP proteins. In this aspect, YCH337 behaved differently from the combination of vincristine and etoposide. YCH337 inhibited proliferation of tumor cells with an averaged IC50 of 0.3 µM. It significantly suppressed the growth of HT-29 xenografts in nude mice too. Importantly, YCH337 nearly equally killed different-mechanism-mediated resistant tumor cells and corresponding parent cells. Together with the novelty of its chemical structure, YCH337 could serve as a promising lead for drug development and a prototype for a dual microtubule/Top2 targeting strategy for cancer therapy.


Asunto(s)
Carbolinas/uso terapéutico , Roturas del ADN de Doble Cadena/efectos de los fármacos , ADN de Neoplasias/efectos de los fármacos , Proteínas de Neoplasias/antagonistas & inhibidores , Inhibidores de Topoisomerasa II/uso terapéutico , Moduladores de Tubulina/uso terapéutico , Animales , Apoptosis/efectos de los fármacos , Sitios de Unión/efectos de los fármacos , Unión Competitiva , Carbolinas/farmacología , Línea Celular Tumoral , Colchicina/metabolismo , Neoplasias del Colon/tratamiento farmacológico , ADN-Topoisomerasas de Tipo II/fisiología , ADN Superhelicoidal/efectos de los fármacos , Interacciones Farmacológicas , Ensayos de Selección de Medicamentos Antitumorales , Etopósido/farmacología , Humanos , Concentración 50 Inhibidora , Metafase/efectos de los fármacos , Ratones , Ratones Desnudos , Microtúbulos/efectos de los fármacos , Microtúbulos/metabolismo , Estructura Molecular , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/fisiología , Paclitaxel/farmacología , Inhibidores de Topoisomerasa II/síntesis química , Inhibidores de Topoisomerasa II/farmacología , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/síntesis química , Moduladores de Tubulina/farmacología , Vincristina/farmacología
11.
Dalton Trans ; 44(13): 6140-57, 2015 Apr 07.
Artículo en Inglés | MEDLINE | ID: mdl-25736331

RESUMEN

4-(p-X-phenyl)thiosemicarbazone of napthaldehyde {where X = Cl (HL¹) and X = Br (HL²)}, thiosemicarbazone of quinoline-2-carbaldehyde (HL³) and 4-(p-fluorophenyl)thiosemicarbazone of salicylaldehyde (H2L4) and their copper(I) {[Cu(HL¹)(PPh3)2Br]·CH3CN (1) and [Cu(HL²)(PPh3)2Cl]·DMSO (2)} and copper(II) {[(Cu2L³2Cl)2(µ-Cl)2]·2H2O (3) and [Cu(L4)(Py)] (4)} complexes are reported herein. The synthesized ligands and their copper complexes were successfully characterized by elemental analysis, cyclic voltammetry, NMR, ESI-MS, IR and UV-Vis spectroscopy. Molecular structures of all the Cu(I) and Cu(II) complexes have been determined by X-ray crystallography. All the complexes (1-4) were tested for their ability to exhibit DNA-binding and -cleavage activity. The complexes effectively interact with CT-DNA possibly by groove binding mode, with binding constants ranging from 104 to 105 M⁻¹. Among the complexes, 3 shows the highest chemical (60%) as well as photo-induced (80%) DNA cleavage activity against pUC19 DNA. Finally, the in vitro antiproliferative activity of all the complexes was assayed against the HeLa cell line. Some of the complexes have proved to be as active as the clinical referred drugs, and the greater potency of 3 may be correlated with its aqueous solubility and the presence of the quinonoidal group in the thiosemicarbazone ligand coordinated to the metal.


Asunto(s)
Antineoplásicos/síntesis química , Complejos de Coordinación/síntesis química , Cobre/química , División del ADN , Tiosemicarbazonas/química , Antineoplásicos/química , Supervivencia Celular/efectos de los fármacos , Complejos de Coordinación/química , Cristalografía por Rayos X , ADN/efectos de los fármacos , ADN/genética , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/genética , ADN Superhelicoidal/efectos de la radiación , Células HeLa , Humanos , Estructura Molecular , Solubilidad
12.
J Agric Food Chem ; 63(3): 864-71, 2015 Jan 28.
Artículo en Inglés | MEDLINE | ID: mdl-25553507

RESUMEN

Date seed protein hydrolysates were evaluated for antioxidant activity as well as solubility and water-holding capacity in food and biological model systems. Date seed protein hydrolysates as well as carnosine exhibited >80% of solubility over a pH range of 2-12. The hydrolysates and carnosine at 0.5% (w/w) were also found to be effective in enhancing water-holding capacity and cooking yield in a fish model system, which was nearly similar to sodium tripolyphosphate (STPP; 0.3%, w/w). Incorporation of hydrolysates (200 ppm) in fish model systems resulted in the highest inhibition (30%) of oxidation in comparison to butylated hydroxytoluene (BHT; 9%). In addition, hydrolysates and carnosine inhibited ß-carotene oxidation by 75%. The hydrolysates (0.1 mg/mL) inhibited LDL cholesterol oxidation by 60%, whereas carnosine inhibited oxidation by 80% after 12 h of incubation. Additionally, hydrolysates and carnosine effectively inhibited hydroxyl (6 mg/mL) and peroxyl (0.1 mg/mL) radical-induced DNA scission. Therefore, date seed protein hydrolysates could be used as a potential functional food ingredient for health promotion.


Asunto(s)
Antioxidantes/análisis , Carnosina/farmacología , Alimentos , Phoeniceae , Proteínas de Plantas/farmacología , Semillas/química , Animales , Carnosina/química , ADN Superhelicoidal/efectos de los fármacos , Peces , Alimentos Funcionales , Humanos , Peroxidación de Lípido/efectos de los fármacos , Lipoproteínas LDL/química , Proteínas de Plantas/química , Hidrolisados de Proteína/farmacología , Alimentos Marinos , Solubilidad , Agua/metabolismo
13.
Eur J Med Chem ; 89: 266-78, 2015 Jan 07.
Artículo en Inglés | MEDLINE | ID: mdl-25462243

RESUMEN

Metal complexes of the type Mn(bpy)2(N3)2 (1), Co(bpy)2(N3)2·3H2O (2) and Zn2(bpy)2(N3)4 (3) (Where bpy = 2,2-bipyridine) have been synthesized and characterized by elemental analysis and spectral (FT-IR, UV-vis) studies. The structure of complexes (1-3) have been determined by single crystal X-ray diffraction studies and the configuration of ligand-coordinated metal(II) ion was well described as distorted octahedral coordination geometry for Mn(II), Co(II) and distorted square pyramidal geometry for Zn(II) complexes. DNA binding interaction of these complexes (1-3) were investigated by UV-vis absorption, fluorescence circular dichroism spectral and molecular docking studies. The intrinsic binding constants Kb of complexes 1, 2 and 3 with CT-DNA obtained from UV-vis absorption studies were 8.37 × 10(4), 2.23 × 10(5) and 5.52 × 10(4) M(-1) respectively. The results indicated that the three complexes are able to bind to DNA with different binding affinity, in the order 2 > 1 > 3. Complexes (1-3) exhibit a good binding propensity to bovine serum albumin (BSA) proteins having relatively high binding constant values. Gel electrophoresis assay demonstrated the ability of the complexes 1-3 promote the cleavage ability of the pBR322 plasmid DNA in the presence of the reducing agent 3-mercaptopropionic acid (MPA) but with different cleavage mechanisms: the complex 3 cleaves DNA via hydrolytic pathway (T4 DNA ligase assay), while the DNA cleavage by complexes 1 and 2 follows oxidative pathway. The chemical nuclease activity follows the order: 2 > 1 > 3. The effects of various activators were also investigated and the nuclease activity efficacy followed the order MPA > GSH > H2O2 > Asc. The cytotoxicity studies of complexes 1-3 were tested in vitro on breast cancer cell line (MCF-7) and they found to be active.


Asunto(s)
2,2'-Dipiridil/química , Antineoplásicos/síntesis química , Azidas/química , Cobalto/química , Complejos de Coordinación/síntesis química , Manganeso/química , Zinc/química , Antineoplásicos/química , Antineoplásicos/farmacología , Supervivencia Celular/efectos de los fármacos , Complejos de Coordinación/química , Complejos de Coordinación/farmacología , División del ADN/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Descubrimiento de Drogas , Células MCF-7 , Simulación del Acoplamiento Molecular , Unión Proteica , Albúmina Sérica Bovina/metabolismo , Difracción de Rayos X
14.
Indian J Exp Biol ; 52(10): 952-64, 2014 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-25345244

RESUMEN

In vitro assessment showed that H. rhamnoides (HrLE) extract possessed free radical scavenging activities and can protect gamma (gamma) radiation induced supercoiled DNA damage. For in vivo study, Swiss albino mice were administered with HrLE (30 mg/kg body weight) for 15 consecutive days before exposing them to a single dose of 5 Gy of beta radiation. HrLE significantly prevented the radiation induced genomic DNA damage indicated as a significant reduction in the comet parameters. The lipid peroxidation, liver function enzymes, expression of phosphorylated NFkappaB (p65) and IkappaBalpha increased whereas the endogenous antioxidants diminished upon radiation exposure compared to control. Pretreatment of HrLE extract ameliorated these changes. Based on the present results it can be concluded that H. rhamnoides possess a potential preventive element in planned and accidental nuclear exposures.


Asunto(s)
Daño del ADN/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Hippophae/química , Hígado/efectos de los fármacos , Extractos Vegetales/farmacología , Animales , ADN Superhelicoidal/química , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/efectos de la radiación , Depuradores de Radicales Libres/química , Rayos gamma , Hígado/química , Hígado/patología , Masculino , Ratones , Extractos Vegetales/química , Hojas de la Planta/química
15.
Bioorg Med Chem Lett ; 24(17): 4215-22, 2014 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-25086682

RESUMEN

A series of dual-targeting, alcohol-containing benzothiazoles has been identified with superior antibacterial activity and drug-like properties. Early lead benzothiazoles containing carboxylic acid moieties showed efficacy in a well-established in vivo model, but inferior drug-like properties demanded modifications of functionality capable of demonstrating superior efficacy. Eliminating the acid group in favor of hydrophilic alcohol moieties at C(5), as well as incorporating solubilizing groups at the C(7) position of the core ring provided potent, broad-spectrum Gram-positive antibacterial activity, lower protein binding, and markedly improved efficacy in vivo.


Asunto(s)
Antibacterianos/farmacología , Benzotiazoles/química , Benzotiazoles/farmacología , ADN Bacteriano/química , ADN Bacteriano/efectos de los fármacos , ADN Superhelicoidal/efectos de los fármacos , Haemophilus influenzae/efectos de los fármacos , Alcoholes/química , Antibacterianos/síntesis química , Antibacterianos/química , Benzotiazoles/síntesis química , Relación Dosis-Respuesta a Droga , Descubrimiento de Drogas , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Staphylococcus , Relación Estructura-Actividad
16.
BMC Genomics ; 15: 291, 2014 Apr 16.
Artículo en Inglés | MEDLINE | ID: mdl-24734910

RESUMEN

BACKGROUND: It has been shown previously that aminocoumarin antibiotics such as novobiocin lead to immediate downregulation of recA expression and thereby inhibit the SOS response, mutation frequency and recombination capacity in Staphylococcus aureus. Aminocoumarins function by inhibiting the ATPase activity of DNA gyrase subunit B with a severe impact on DNA supercoiling. RESULTS: Here, we have analysed the global impact of the DNA relaxing agent novobiocin on gene expression in S. aureus. Using a novobiocin-resistant mutant, it became evident that the change in recA expression is due to gyrase inhibition. Microarray analysis and northern blot hybridisation revealed that the expression levels of a distinct set of genes were increased (e.g., recF-gyrB-gyrA, the rib operon and the ure operon) or decreased (e.g., arlRS, recA, lukA, hlgC and fnbA) by novobiocin. The two-component ArlRS system was previously found to decrease the level of supercoiling in S. aureus. Thus, downregulation of arlRS might partially compensate for the relaxing effect of novobiocin. Global analysis and gene mapping of supercoiling-sensitive genes did not provide any indication that they are clustered in the genome. Promoter fusion assays confirmed that the responsiveness of a given gene is intrinsic to the promoter region but independent of the chromosomal location. CONCLUSIONS: The results indicate that the molecular properties of a given promoter, rather than the chromosomal topology, dictate the responsiveness to changes in supercoiling in the pathogen Staphylococcus aureus.


Asunto(s)
Aminocumarinas/farmacología , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genética , Antineoplásicos/farmacología , Proteínas Bacterianas/genética , Girasa de ADN/metabolismo , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/genética , Perfilación de la Expresión Génica , Genoma Bacteriano , Familia de Multigenes , Regiones Promotoras Genéticas , Rec A Recombinasas/genética , Rec A Recombinasas/metabolismo , Virulencia/genética
17.
Chem Biol Interact ; 217: 9-18, 2014 Jun 25.
Artículo en Inglés | MEDLINE | ID: mdl-24732435

RESUMEN

The combination of cisplatin and ionizing radiation (IR) increases cell toxicity by both enhancing DNA damage and inhibiting repair mechanisms. Although the formation of cluster DNA lesions, particularly double-strand breaks (DSB) at the site of cisplatin-DNA-adducts has been reported to induce cell death, the contribution of DSB and non-DSB cluster lesions to the cellular toxicity is still unknown. Although both lesions are toxic, it is not always possible to measure their frequency and cell survival in the same model system. To overcome this problem, here, we investigate the effect of cisplatin-adducts on the induction of DSB and non-DSB cluster DNA lesions by IR and determine the impact of such lesions on plasmid functionality. Cluster lesions are two or more lesions on opposite DNA strands with a short distance such that error free repair is difficult or impossible. At a ratio of two cisplatin per plasmid, irradiation of platinated DNA in solution with (137)Cs γ-rays shows enhancements in the formation of DNA DSB and non-DSB cluster lesions by factors of 2.6 and 2.1, respectively, compared to unmodified DNA. However, in absolute terms, the yield for non-DSB cluster lesions is far larger than that for DSB, by a factor of 26. Unmodified and cisplatin-modified DNA were irradiated and subsequently transformed into Escherichia coli to give survival curves representing the functionality of the plasmid DNA as a function of radiation dose. Our results demonstrate that non-DSB cluster lesions are the only toxic lesions present at a sufficient frequency to account for the loss of DNA functionality. Our data also show that Frank-DSB lesions are simply too infrequent to account for the loss of DNA functionality. In conclusion, non-DSB cluster DNA damage is known to be difficult to repair and is probably the lesion responsible for the loss of functionality of DNA modified by cisplatin.


Asunto(s)
Cisplatino/metabolismo , Cisplatino/efectos de la radiación , Aductos de ADN/metabolismo , Aductos de ADN/efectos de la radiación , Daño del ADN , Reparación del ADN , ADN/efectos de los fármacos , ADN/efectos de la radiación , Plásmidos/efectos de la radiación , Cisplatino/química , Cisplatino/farmacología , ADN/metabolismo , ADN Superhelicoidal/efectos de los fármacos , ADN Superhelicoidal/metabolismo , ADN Superhelicoidal/efectos de la radiación , Rayos gamma , Radiación Ionizante
18.
J Antimicrob Chemother ; 69(8): 2102-9, 2014 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-24729602

RESUMEN

OBJECTIVES: Loop B is important for low-level quinolone resistance conferred by Qnr proteins. The role of individual amino acids within QnrS1 loop B in quinolone resistance and gyrase protection was assessed. METHODS: qnrS1 and 11 qnrS1 alleles with site-directed Ala mutations in loop B were expressed in Escherichia coli BL21(DE3) and proteins were purified by affinity chromatography. Ciprofloxacin MICs were determined with and without IPTG. Gyrase DNA supercoiling was measured with and without ciprofloxacin IC50 and with various concentrations of QnrS1 proteins. RESULTS: Wild-type QnrS1 and QnrS1 with Asn-110→Ala and Arg-111→Ala substitutions increased the ciprofloxacin MIC 12-fold in BL21(DE3), although QnrS1 with Gln-107→Ala replacement increased it 2-fold more than wild-type did. However, QnrS1 with Ala substitutions at His-106, Val-108, Ser-109, Met-112, Tyr-113, Phe-114, Cys-115 and Ser-116 increased ciprofloxacin MIC 1.4- to 8-fold less than wild-type QnrS1. Induction by 10-1000 µM IPTG increased ciprofloxacin MICs for all mutants, reaching values similar to those for wild-type. Purified wild-type and mutated proteins differed in protection of gyrase from ciprofloxacin action. Wild-type QnrS1 produced complete protection of gyrase supercoiling from ciprofloxacin (1.8 µM) action at 0.05 nM and half protection at 0.5 pM, whereas QnrS1 with Ala replacements that conferred the least increase in ciprofloxacin MICs also required the highest QnrS1 concentrations for protection. CONCLUSIONS: Key individual residues in QnrS1 loop B affect ciprofloxacin resistance and gyrase protection from ciprofloxacin action, supporting the concept that loop B is key for interaction with gyrase necessary for quinolone resistance.


Asunto(s)
Ciprofloxacina/farmacología , Farmacorresistencia Bacteriana/genética , Proteínas de Escherichia coli/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Sustitución de Aminoácidos , Antibacterianos/farmacología , Girasa de ADN/efectos de los fármacos , Girasa de ADN/genética , ADN Superhelicoidal/efectos de los fármacos , Escherichia coli/metabolismo , Isopropil Tiogalactósido/farmacología , Pruebas de Sensibilidad Microbiana , Mutación , Estructura Terciaria de Proteína , Relación Estructura-Actividad
19.
Exp Parasitol ; 140: 24-32, 2014 May.
Artículo en Inglés | MEDLINE | ID: mdl-24594261

RESUMEN

2-Cys peroxiredoxin (Prx) is the main antioxidant enzyme in Fasciola species for detoxifying hydrogen peroxide which is generated from the hosts' immune effector cells and the parasites' own metabolism. In this study, the recombinant Prx protein from Fasciola gigantica (rFgPrx-2) was expressed and purified in a prokaryotic expression system. This recombinant protein with molecular weight of 26 kDa was enzymatically active in reduction of hydrogen peroxide both in presence of thioredoxin and glutathione systems, and also protected the supercoiled plasmid DNA from oxidative damage in metal-catalyzed oxidation (MCO) system in a concentration-dependent manner. By immunoblotting, using antibody against rFgPrx-2 as probe, a native FgPrxs, whose MW at 25 kDa, was detected in all developmental stages of the parasite. Concentrations of native FgPrxs were increasing in all stages reaching highest level in adult stage. The antibody also showed cross reactivities with corresponding proteins in some cattle helminthes. Natural antibody to FgPrxs could be detected in the sera of mice at 3 and 4 weeks after infection with F. gigantica metacercariae. By immunofluorescence, FgPrxs was highly expressed in tegument and tegumental cells, parenchyma, moderately expressed in cecal epithelial cells in early, juvenile and adult worms. Furthermore, FgPrxs was also detected in the female reproductive organs, including eggs, ovary, vitelline cells, and testis, suggesting that FgPrxs might play an essential role in protecting parasite's tissues from free radical attack during their life cycle. Thus, FgPrxs is one potential candidate for drug therapy and vaccine development.


Asunto(s)
Antioxidantes/metabolismo , Fasciola/metabolismo , Proteínas del Helminto/metabolismo , Peróxido de Hidrógeno/metabolismo , Peroxirredoxinas/metabolismo , Animales , Antioxidantes/química , Bovinos , Daño del ADN , ADN Superhelicoidal/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Fasciola/genética , Fasciola/inmunología , Femenino , Regulación de la Expresión Génica , Glutatión/metabolismo , Proteínas del Helminto/química , Proteínas del Helminto/genética , Ratones , Ratones Endogámicos ICR , Peso Molecular , Oxidación-Reducción , Peroxirredoxinas/química , Peroxirredoxinas/genética , Plásmidos , Conejos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tiorredoxinas/metabolismo
20.
Biochem Biophys Res Commun ; 444(1): 50-5, 2014 Jan 31.
Artículo en Inglés | MEDLINE | ID: mdl-24434150

RESUMEN

The intercalating drugs possess a planar aromatic chromophore unit by which they insert between DNA bases causing the distortion of classical B-DNA form. The planar tricyclic structure of anthraquinones belongs to the group of chromophore units and enables anthraquinones to bind to DNA by intercalating mode. The interactions of simple derivatives of anthraquinone, quinizarin (1,4-dihydroxyanthraquinone) and danthron (1,8-dihydroxyanthraquinone), with negatively supercoiled and linear DNA were investigated using a combination of the electrophoretic methods, fluorescence spectrophotometry and single molecule technique an atomic force microscopy. The detection of the topological change of negatively supercoiled plasmid DNA, unwinding of negatively supercoiled DNA, corresponding to appearance of DNA topoisomers with the low superhelicity and an increase of the contour length of linear DNA in the presence of quinizarin and danthron indicate the binding of both anthraquinones to DNA by intercalating mode.


Asunto(s)
Antraquinonas/farmacología , ADN Superhelicoidal/química , ADN Superhelicoidal/efectos de los fármacos , ADN/química , ADN/efectos de los fármacos , ADN Forma B/química , ADN Forma B/efectos de los fármacos , Electroforesis en Gel de Gradiente Desnaturalizante , Sustancias Intercalantes/farmacología , Microscopía de Fuerza Atómica , Conformación de Ácido Nucleico/efectos de los fármacos , Plásmidos/química , Plásmidos/efectos de los fármacos , Espectrometría de Fluorescencia
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