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1.
Nucleic Acids Res ; 48(15): 8675-8685, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32687167

RESUMO

Along with nucleobase pairing, base-base stacking interactions are one of the two main types of strong non-covalent interactions that define the unique secondary and tertiary structure of RNA. In this paper we studied two subfamilies of nucleobase-inserted stacking structures: (i) with any base intercalated between neighboring nucleotide residues (base-intercalated element, BIE, i + 1); (ii) with any base wedged into a hydrophobic cavity formed by heterocyclic bases of two nucleotides which are one nucleotide apart in sequence (base-wedged element, BWE, i + 2). We have exploited the growing database of natively folded RNA structures in Protein Data Bank to analyze the distribution and structural role of these motifs in RNA. We found that these structural elements initially found in yeast tRNAPhe are quite widespread among the tertiary structures of various RNAs. These motifs perform diverse roles in RNA 3D structure formation and its maintenance. They contribute to the folding of RNA bulges and loops and participate in long-range interactions of single-stranded stretches within RNA macromolecules. Furthermore, both base-intercalated and base-wedged motifs participate directly or indirectly in the formation of RNA functional centers, which interact with various ligands, antibiotics and proteins.


Assuntos
Complexos Multiproteicos/ultraestrutura , Conformação de Ácido Nucleico , Proteínas de Ligação a RNA/ultraestrutura , RNA/ultraestrutura , Antibacterianos/química , Pareamento de Bases/genética , Substâncias Intercalantes/química , Ligantes , Modelos Moleculares , Conformação Molecular , Complexos Multiproteicos/química , Complexos Multiproteicos/genética , Nucleotídeos/química , Nucleotídeos/genética , RNA/química , RNA/genética , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética
2.
Eur J Med Chem ; 200: 112407, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32512480

RESUMO

The photophysical properties of naphthalimide-based fluorophores can be easily tuned by chemical manipulation of the substituents on that privileged scaffold. Replacement of a OMe group at position 6 in 2-(hydroxyl)ethyl-naphthalimide derivatives by diverse amines, including 2-(hydroxyl)ethylamine, trans-(4-acetamido)cyclohexylamine and azetidine increases the solvatochromic (ICT) character, while this replacement in 2-(dimethylamino)ethyl-naphthalimide analogues (PET fluorophores) decrease their solvent polarity sensitivity or even reversed them to solvatochromic fluorophores. These fluorophores resulted macrophage nucleus imaging probes, which bind DNA as intercalants and showed low cytotoxicity in human cancer cells.


Assuntos
Núcleo Celular , Corantes Fluorescentes/química , Macrófagos/ultraestrutura , Imagem Molecular/métodos , Naftalimidas/química , Linhagem Celular Tumoral , DNA/metabolismo , Humanos , Substâncias Intercalantes/química , Relação Estrutura-Atividade
3.
Adv Exp Med Biol ; 1195: 59-71, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32468460

RESUMO

Herein, we deploy an in silico pipeline of structural bioinformatics, thermodynamics, and molecular dynamics to investigate the role of cortisol in circadian rhythms, biorhythms, stress response, and even sleep disorders. Our study shows that high concentrations of cortisol intercalate in the minor groove of DNA. This phenomenon widens the adjacent major grooves and provides the Clock/Bmal1 complex with more space to dock and interact with DNA. Then, the strong charges of cortisol pull the alpha helices of the Clock/Bmal1 complex and bend it inward, thus establishing stronger interactions and prolonged signaling. Our results indicate that elevated cortisol levels play an important role in stress, inflammation, and sleep disorders as a result of prolonged and stronger dsDNA - Clock/Bmal1 interactions.


Assuntos
Fatores de Transcrição ARNTL/metabolismo , Proteínas CLOCK/metabolismo , DNA/química , Hidrocortisona/metabolismo , Sistema Hipotálamo-Hipofisário/fisiologia , Sistema Hipófise-Suprarrenal/fisiologia , Transtornos do Sono-Vigília/fisiopatologia , Estresse Psicológico/prevenção & controle , Ritmo Circadiano/genética , Ritmo Circadiano/fisiologia , Simulação por Computador , DNA/metabolismo , Humanos , Hidrocortisona/química , Inflamação/genética , Inflamação/metabolismo , Substâncias Intercalantes/química , Substâncias Intercalantes/metabolismo , Receptores de Glucocorticoides/metabolismo , Transtornos do Sono-Vigília/genética , Estresse Psicológico/genética , Estresse Psicológico/fisiopatologia
4.
J Phys Chem Lett ; 11(7): 2436-2442, 2020 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-32141760

RESUMO

Herein we report the effect of different nucleobase pair compositions on the association-induced fluorescence enhancement property of Thioflavin T (ThT), upon binding with 20 base pair long double-stranded DNA (dsDNA). Analysis of binding and decay constants along with the association (Kass) and dissociation (Kdiss) rate constants obtained from the fluctuation in the fluorescence intensity of ThT after binding with different DNA revealed selective affinity of ThT toward AT-rich dsDNA. Molecular docking also substantiates the experimental results. We also observed that addition of orange-emitting ethidium bromide (EtBr) to cyan-emitting ThT-DNA complexes leads to bright white light emission (WLE) through Förster resonance energy transfer. Additionally, the emission of white light is far greater in the case of intra-DNA strands. Besides endorsing the binding insights of ThT to AT-rich dsDNA, the present investigations open a new perspective for realizing promising WLE from two biomarkers without labeling the DNA.


Assuntos
Benzotiazóis/metabolismo , DNA/metabolismo , Substâncias Intercalantes/metabolismo , Benzotiazóis/química , DNA/química , Etídio/química , Etídio/metabolismo , Transferência Ressonante de Energia de Fluorescência , Substâncias Intercalantes/química , Luz
5.
Nanotechnology ; 31(23): 235605, 2020 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-32125281

RESUMO

Intercalation of drug molecules into synthetic DNA nanostructures formed through self-assembled origami has been postulated as a valuable future method for targeted drug delivery. This is due to the excellent biocompatibility of synthetic DNA nanostructures, and high potential for flexible programmability including facile drug release into or near to target cells. Such favourable properties may enable high initial loading and efficient release for a predictable number of drug molecules per nanostructure carrier, important for efficient delivery of safe and effective drug doses to minimise non-specific release away from target cells. However, basic questions remain as to how intercalation-mediated loading depends on the DNA carrier structure. Here we use the interaction of dyes YOYO-1 and acridine orange with a tightly-packed 2D DNA origami tile as a simple model system to investigate intercalation-mediated loading. We employed multiple biophysical techniques including single-molecule fluorescence microscopy, atomic force microscopy, gel electrophoresis and controllable damage using low temperature plasma on synthetic DNA origami samples. Our results indicate that not all potential DNA binding sites are accessible for dye intercalation, which has implications for future DNA nanostructures designed for targeted drug delivery.


Assuntos
Laranja de Acridina/química , Benzoxazóis/química , DNA/química , Substâncias Intercalantes/química , Compostos de Quinolínio/química , Sítios de Ligação , Eletroforese em Gel Bidimensional , Microscopia de Força Atômica , Microscopia de Fluorescência , Modelos Moleculares , Nanoestruturas/química , Conformação de Ácido Nucleico , Imagem Individual de Molécula
6.
Org Biomol Chem ; 18(8): 1671-1678, 2020 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-32051993

RESUMO

The dynamic nature of nucleic acid alkylation by simple ortho quinone methides (QM) and their conjugates has provided numerous opportunities ranging from sequence selective targeting to bipedal walking in duplex DNA. To enhance the diffusion rate of adduct migration, one of two sites for QM generation was deleted from a bisQM conjugate of acridine to remove the covalent anchor to DNA that persists during QM regeneration. This conversion of a bisfunctional cross-linking agent to a monofunctional alkylating agent allowed adduct diffusion to traverse an extrahelical -TT- bulge that previously acted as a barrier for its bisfunctional analog. An electron rich derivative of the monofunctional acridine conjugate was additionally prepared to accelerate the rates of DNA alkylation and QM regeneration. The resulting stabilization of this QM effectively enhanced the rate of its release from adducts attached at guanine N7 in competition with an alternative and detrimental deglycosylation pathway. Intercalation by the acridine component was not sufficient to hold the transient QM intermediates within duplex DNA and consequently these electrophiles diffused into solution and were subject to quenching by solvent and a model nucleophile, ß-mercaptoethanol.


Assuntos
Acridinas/química , DNA/metabolismo , Indolquinonas/química , Alquilantes/química , Alquilação , DNA/química , Adutos de DNA/química , Difusão , Substâncias Intercalantes/química
7.
Chem Commun (Camb) ; 56(13): 1996-1999, 2020 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-31960843

RESUMO

Here, we developed a coralyne-based, 'light-up' intercalator displacement assay to identify molecular stabilizers of triplex DNA using a sequence from a chromosomal breakpoint hotspot in the human c-MYC oncogene. Its potential to identify triplex DNA ligands was demonstrated using BePI and doxorubicin. Identification of triplex-interacting ligands may allow the regulation of genetic instability in human genomes.


Assuntos
DNA/análise , Substâncias Intercalantes/química , Alcaloides de Berberina/química , DNA/química , Doxorrubicina/química , Genoma Humano , Instabilidade Genômica , Humanos , Indóis/química , Ligantes , Proteínas Proto-Oncogênicas c-myc/genética , Piridinas/química
8.
Chemistry ; 26(1): 259-268, 2020 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-31614021

RESUMO

In the effort to overcome issues of toxicity and resistance inherent to treatment by the approved platinum anticancer agents, a large number of cisplatin variants continues today to be prepared and tested. One of the applied strategies is to use monofunctional platinum complexes that, unlike traditional bifunctional compounds, are able to form only a single covalent bond with nuclear DNA. Chirality, aquation reaction, interaction with guanine and N-acetyl methionine as well as, intercalation into, binding to and distortion of DNA have been investigated by using both quantum mechanical DFT and molecular dynamics computations aiming at contributing to the elucidation of the molecular mechanism underlying the significantly enhanced spectrum of activity of the monofunctional PtII drug phenanthriplatin. Analogous calculations have been performed in parallel for other two less potent monofunctional PtII drugs, pyriplatin and enpyriplatin, which show very different cytotoxic effects.


Assuntos
Antineoplásicos/química , Teoria da Densidade Funcional , Simulação de Dinâmica Molecular , Compostos Organoplatínicos/química , Fenantridinas/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , DNA/química , DNA/metabolismo , Humanos , Substâncias Intercalantes/química , Metionina/química , Conformação de Ácido Nucleico , Compostos Organoplatínicos/farmacologia , Fenantridinas/farmacologia , Termodinâmica , Transcrição Genética/efeitos dos fármacos
9.
Talanta ; 206: 120217, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514897

RESUMO

MicroRNAs (miRNA) involve in regulating different physiological processes whose dysregulation is associated with a wide range of diseases including cancers, diabetes and cardiovascular problems. Herein, we report a direct, sensitive and highly selective detection assay for circulating microRNA (miRNA). This detection strategy employs magnetic nanoparticles as the reaction platform which can not only allow online pre-concentration and selective separation but also integrates ligation reaction with amplification to enhance the sensitivity of the detection assay. With the presence of the target miRNA, the locked nucleic acid (LNA)-modified molecular beacon (MB) opens up, exposing the binding sites at two ends. The 3'- and 5'-end of the MB responsible for the attachment onto the magnetic nanoparticles, and reporting probe for the attachment of the pair of amplification probes respectively. The ligase ligate RNA to DNA enhance the amplification efficiency. Upon labelled with intercalating fluorophores (YOYO-1) on the hybrids, the quantification of the target miRNA was determined by measuring the fluorescence intensity. A detection limit of 314 fM was achieved with trace amount of sample consumption (~20 µL). As a proof of concept, miRNA-149 was chosen as the target miRNA. This assay is capable of discriminating single-base and reliably quantifying circulating miRNA-149 in both healthy and cancer patient's serums. The result obtained was comparable with that of quantitative reverse transcription polymerase chain reaction (qRT-PCR), suggesting that this direct and sensitive assay can be served as a promising, non-invasive tool for early diagnosis of breast cancer and colorectal cancer.


Assuntos
MicroRNA Circulante/sangue , MicroRNAs/sangue , Microscopia de Fluorescência/métodos , RNA Ligase (ATP)/química , Proteínas Virais/química , Benzoxazóis/química , MicroRNA Circulante/genética , DNA/química , DNA/genética , Corantes Fluorescentes/química , Humanos , Substâncias Intercalantes/química , Limite de Detecção , Nanopartículas de Magnetita/química , MicroRNAs/genética , Hibridização de Ácido Nucleico , Oligonucleotídeos/química , Oligonucleotídeos/genética , Estudo de Prova de Conceito , Compostos de Quinolínio/química
10.
Molecules ; 24(24)2019 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-31835555

RESUMO

A series of ZnCl2 complexes (compounds 1-10) with 4'-(substituted-phenyl)-2,2':6',2''-terpyridine that bears hydrogen (L1), p-methyl (L2), p-methoxy (L3), p-phenyl (L4), p-tolyl (L5), p-hydroxyl (L6), m-hydroxyl (L7), o-hydroxyl (L8), p-carboxyl (L9), or p-methylsulfonyl (L10) were prepared and then characterized by 1H NMR, electrospray mass-spectra (ESI-MS), IR, elemental analysis, and single crystal X-ray diffraction. In vitro cytotoxicity assay was used to monitor the antiproliferative activities against tumor cells. Absorption spectroscopy, fluorescence titration, circular dichroism spectroscopy, and molecular modeling studied the DNA interactions. All of the compounds display interesting photoluminescent properties and different maximal emission peaks due to the difference of the substituent groups. The cell viability studies indicate that the compounds have excellent antiproliferative activity against four human carcinoma cell lines, A549, Bel-7402, MCF-7, and Eca-109, with the lowest IC50 values of 0.33 (10), 0.66 (6), 0.37 (7), and 1.05 (7) µM, respectively. The spectrophotometric results reveal that the compounds have strong affinity binding with DNA as intercalator and induce DNA conformational transition. Molecular docking studies indicate that the binding is contributed by the π…π stacking and hydrogen bonds, providing an order of nucleotide sequence binding selectivity as ATGC > ATAT > GCGC. These compounds intercalate into the base pairs of the DNA of the tumor cells to affect their replication and transcription, and the process is supposed to play an important role in the anticancer mechanism.


Assuntos
Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Piridinas/química , Zinco/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , DNA/química , Relação Dose-Resposta a Droga , Humanos , Substâncias Intercalantes/química , Substâncias Intercalantes/farmacologia , Modelos Moleculares , Relação Estrutura-Atividade
11.
Int J Mol Sci ; 20(24)2019 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-31817267

RESUMO

Ovarian cancer is the most lethal gynecological malignancy, often because of the frequent insurgence of chemoresistance to the drugs currently used. Thus, new therapeutical agents are needed. We tested the toxicity of 16 new DNA-intercalating agents to cisplatin (cDDP)-sensitive human ovarian carcinoma cell lines and their resistant counterparts. The compounds were the complexes of Pt(II) or Pd(II) with bipyridyl (bipy) and phenanthrolyl (phen) and with four different thiourea ancillary ligands. Within each of the four series of complexes characterized by the same thiourea ligand, the Pd(phen) drugs invariably showed the highest anti-proliferative efficacy. This paralleled both a higher intracellular drug accumulation and a more efficient DNA intercalation than all the other metal-bidentate ligand combinations. The consequent inhibition of topoisomerase II activity led to the greatest inhibition of DNA metabolism, evidenced by the inhibition of the expression of the folate cycle enzymes and a marked perturbation of cell-cycle distribution in both cell lines. These findings indicate that the particular interaction of Pd(II) with phenanthroline confers the best pharmacokinetic and pharmacodynamic properties that make this class of DNA intercalators remarkable inhibitors, even of the resistant cell growth.


Assuntos
Proliferação de Células/efeitos dos fármacos , Complexos de Coordenação/química , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Substâncias Intercalantes/farmacologia , Fenantrolinas/química , Tioureia/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cisplatino/farmacologia , Complexos de Coordenação/metabolismo , Complexos de Coordenação/farmacologia , DNA/química , DNA/metabolismo , DNA Topoisomerases Tipo II/química , DNA Topoisomerases Tipo II/metabolismo , Feminino , Humanos , Substâncias Intercalantes/química , Ligantes , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Paládio/química , Platina/química
12.
Mikrochim Acta ; 186(12): 826, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31754803

RESUMO

A photoelectrochemical (PEC) aptasensor for Pb(II) detection is described. A nanocomposite consisting of CdS (2.5 µm) and TiO2 nanoparticles (10 nm) was used as a photoactive material, and gold nanochains (Au NCs) as the support for immobilization of the Pb(II)-binding aptamer. The quercetin-copper(II) complex was further employed as the intercalator for the improvement of the photoactivity by embedding it into dsDNA. In the presence of Pb(II), a Pb(II)-stabilized G-quadruplex was formed between Pb(II) and DNA S1. This is accompanied by unwinding of the dsDNA and the release of the quercetin-copper(II) complex from the surface of the sensor. This results in a decrease of the photocurrent that drops linearly from 5.0 × 10-12 to 1.0 × 10-8 mol·L-1 Pb(II) concentration range with a detection limit of 1.6 × 10-12 mol·L-1. The method was applied to the determination of Pb(II) in various samples and gave satisfactory results. Graphical abstractA photoelectrochemical aptasensor was fabricated for the detection of Pb(II) based on CdS-TiO2 nanocomposite modified indium tin oxide (ITO) electrode. Gold nanochains (AuNCs) were used as anchor to immobilize the aptamers S1 and S2 that form a double helix structure by DNA hybridization. After embedding of quercetin-copper(II) complex as intercalator and electron donor, the concentrations of Pb(II) were determined by the changes of photocurrents.


Assuntos
Aptâmeros de Nucleotídeos/química , Complexos de Coordenação/química , DNA/química , Substâncias Intercalantes/química , Chumbo/análise , Nanopartículas Metálicas/química , Técnicas Biossensoriais , Compostos de Cádmio/química , Cobre/química , Condutividade Elétrica , Técnicas Eletroquímicas , Eletrodos , Quadruplex G , Ouro/química , Limite de Detecção , Hibridização de Ácido Nucleico , Processos Fotoquímicos , Quercetina/química , Sulfetos/química , Propriedades de Superfície , Titânio/química
13.
Mikrochim Acta ; 186(12): 824, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31754805

RESUMO

The authors describe a tetrahedral DNA nanostructure loaded with SYBR Green (SG-TDN) for fluorometric determination of nucleic acids. After intercalating into the TDN, fluorescence of SG is enhanced by 260-fold (exc 480 nm, em 524 nm), and the resulting SG-TDN nanoflare displays >7-fold stronger fluorescence than that of FAM-labeled TDN. The SG-TDNs were coupled to magnetic microparticles and polydopamine nanoparticles to construct multi-functional nanoprobes through sequence hybridization using a toehold strand. The method was applied to detect a stretch of microRNA sequence (20 bp) in buffer and in undiluted serum with excellent selectivity, over a wide linear range and with a low limit of detection (0.2 nM). The probe was also applied for visualization of tumor-related microRNA in living cells via fluorescence imaging. Graphical abstract Schematic representation of tetrahedron-based DNA nanoflare for fluorometric nucleic acid determination in undiluted blood serum and living cells.


Assuntos
Corantes Fluorescentes/química , Substâncias Intercalantes/química , Nanoestruturas/química , Ácidos Nucleicos/química , Células A549 , Técnicas Biossensoriais , DNA/química , Fluorometria , Células HEK293 , Humanos , Indóis , Limite de Detecção , MicroRNAs/química , Hibridização de Ácido Nucleico , Imagem Óptica , Polímeros , Sensibilidade e Especificidade
14.
J Chem Phys ; 151(16): 164902, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31675856

RESUMO

Most of the anticancer drugs bind to double-stranded DNA (dsDNA) by intercalative-binding mode. Although experimental studies have become available recently, a molecular-level understanding of the interactions between the drug and dsDNA that lead to the stability of the intercalated drug is lacking. Of particular interest are the modifications of the mechanical properties of dsDNA observed in experiments. The latter could affect many biological functions, such as DNA transcription and replication. Here, we probe, via all-atom molecular dynamics (MD) simulations, the change in the mechanical properties of intercalated drug-DNA complexes for two intercalators, daunomycin and ethidium. We find that, upon drug intercalation, the stretch modulus of DNA increases significantly, whereas its persistence length and bending modulus decrease. Steered MD simulations reveal that it requires higher forces to stretch the intercalated dsDNA complexes than the normal dsDNA. Adopting various pulling protocols to study force-induced DNA melting, we find that the dissociation of dsDNA becomes difficult in the presence of intercalators. The results obtained here provide a plausible mechanism of function of the anticancer drugs, i.e., via altering the mechanical properties of DNA. We also discuss long-time consequences of using these drugs, which require further in vivo investigations.


Assuntos
Antineoplásicos/química , DNA/química , Substâncias Intercalantes/química , Simulação de Dinâmica Molecular , Conformação de Ácido Nucleico
15.
Analyst ; 144(23): 6936-6943, 2019 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-31617512

RESUMO

MicroRNAs (miRNAs) are small non-coding RNA molecules that serve as important biomarkers for a variety of diseases such as cancer and vascular disease. However, sensitive and accurate detection of miR-21 is very challenging in that up-regulation of miR-21 is highly associated with several types of malignant tumors. Here, quartz crystal microbalance (QCM) biosensors were developed for sensitive and specific detection of miR-21 through formation of miR-21-DNA hybrid duplexes and non-specific intercalation of surface-modified pyrene molecules. High selectivity for miR-21 over other miRNAs came from the specific hybridization between miR-21 and gold nanoparticle (AuNP)-conjugated complementary oligonucleotides of miR-21. High sensitivity was obtained through formation of intercalated complexes on the surface with subsequent gold staining signal amplification. Under optimum condition using this strategic approach, our novel QCM biosensors could detect miR-21 concentration as low as 3.6 pM in the entire linear range from 2.5 pM to 2.5 µM with a correlation coefficient of 0.989. In addition, these sensors did not work at all for other miRNAs based on their high selectivity. miR-21 in human brain total RNA and total RNA extracted from A549 cell line could also be successfully detected. Therefore, miRNA detection technology using QCM biosensors and their detection mechanisms have potential as alternatives in biological studies and clinical diagnosis.


Assuntos
DNA/química , Substâncias Intercalantes/química , MicroRNAs/análise , Pirenos/química , Células A549 , Técnicas Biossensoriais/métodos , Química Encefálica , DNA/genética , Ouro/química , Humanos , Limite de Detecção , Nanopartículas Metálicas/química , MicroRNAs/genética , Hibridização de Ácido Nucleico , Oligodesoxirribonucleotídeos/química , Oligodesoxirribonucleotídeos/genética , Técnicas de Microbalança de Cristal de Quartzo/métodos
16.
Mikrochim Acta ; 186(11): 707, 2019 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-31637526

RESUMO

A method is described for the determination of DNA via nucleic acid amplification by using nucleic acid concatemers that result from DNA supersandwich self-assemblies (SSAs). The method employs two auxiliary probes to form self-assembled biotin SSAs. These exhibit strong fluorescence if labeled with intercalator SYBR Green I. In the presence of the target (as exemplified for a 30-mer), streptavidin is released from the surface of the functionalized magnetic microparticles (FMMPs) by competitive hybridization on the surface. However, the SSA products do not conjugate to the FMMPs. This leads to a large amount of SYBR Green I intercalated into the concatemers and eventually results in amplified fluorescence in the supernate. The SSA products can be prepared beforehand, and amplification therefore can be completed within 50 min. The method is more efficient than any other conventional amplification. The detection limit for the 30-mer is 26.4 fM which is better by a factor of 10 compared to other amplification methods. Conceivably, the method can be further extended to the determination of a wide variety of targets simply by replacing the sequences of the probes. Finally, this rapid and highly sensitive method was employed for detection of Ebola virus gene (≈30-mer) and ATP in spiked serum with satisfactory results. Graphical abstract A high sensitivity and efficiency bioassay is described based on functionalized magnetic microparticles and DNA supersandwich self-assemblies.


Assuntos
DNA Concatenado/química , DNA de Cadeia Simples/sangue , Fluorometria/métodos , Trifosfato de Adenosina/sangue , Trifosfato de Adenosina/química , Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/genética , Biotina/química , Sondas de DNA/química , Sondas de DNA/genética , DNA Concatenado/genética , DNA de Cadeia Simples/genética , DNA Viral/sangue , DNA Viral/genética , Ebolavirus/química , Humanos , Substâncias Intercalantes/química , Limite de Detecção , Fenômenos Magnéticos , Hibridização de Ácido Nucleico , Compostos Orgânicos/química , Estreptavidina/química
17.
Eur Phys J E Soft Matter ; 42(10): 130, 2019 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-31583481

RESUMO

In this work we use single molecule force spectroscopy performed with optical tweezers in order to characterize the complexes formed between the anticancer drug Pixantrone (PIX) and the DNA molecule, at two very different ionic strengths. Firstly, the changes of the mechanical properties of the DNA-PIX complexes were studied as a function of the drug concentration in the sample. Then, a quenched-disorder statistical model of ligand binding was used in order to determine the physicochemical (binding) parameters of the DNA-PIX interaction. In particular, we have found that the PIX molecular mechanism of action involves intercalation into the double helix, followed by a significant compaction of the DNA molecule due to partial neutralization of the phosphate backbone. Finally, this scenario of interaction was quantitatively compared to that found for the related drug Mitoxantrone (MTX), which binds to DNA with a considerably higher equilibrium binding constant and promotes a much stronger DNA compaction. The comparison performed between the two drugs can bring clues to the development of new (and more efficient) related compounds.


Assuntos
Antineoplásicos/química , DNA/química , Substâncias Intercalantes/química , Isoquinolinas/química , Ligantes , Pinças Ópticas , Imagem Individual de Molécula/métodos
18.
Phys Chem Chem Phys ; 21(38): 21549-21560, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31536074

RESUMO

Repetitive cytosine rich i-motif forming sequences are abundant in the telomere, centromere and promoters of several oncogenes and in some instances are known to regulate transcription and gene expression. The in vivo existence of i-motif structures demands further insight into the factors affecting their formation and stability and development of better understanding of their gene regulatory functions. Most prior studies characterizing the conformational dynamics of i-motifs are based on i-motif forming synthetic constructs. Here, we present a systematic study on the stability and structural properties of biologically relevant i-motifs of telomeric and centromeric repeat fragments. Our results based on molecular dynamics simulations and quantum chemical calculations indicate that along with base pairing interactions within the i-motif core the overall folded conformation is associated with the stable C-HO sugar "zippers" in the narrow grooves and structured water molecules along the wide grooves. The stacked geometry of the hemi-protonated cytosine pairs within the i-motif core is mainly governed by the repulsive base stacking interaction. The loop sequence can affect the structural dynamics of the i-motif by altering the loop motion and backbone conformation. Overall this study provides microscopic insight into the i-motif structure that will be helpful to understand the structural aspect of mechanisms of gene regulation by i-motif DNA.


Assuntos
DNA/química , Substâncias Intercalantes/química , Motivos de Nucleotídeos , Solventes/química , Telômero/química , Pareamento de Bases , Citosina/química , Ligação de Hidrogênio , Simulação de Dinâmica Molecular
19.
Inorg Chem ; 58(19): 13150-13160, 2019 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-31539237

RESUMO

Mitochondrial DNA (mtDNA) is an attractive cellular target for anticancer agents in addition to nuclear DNA (nDNA). The cationic platinum(II) complex cis-[Pt(NP)(NH3)2Cl]NO3 (PtNP, NP = N-(2-ethylpyridine)-1,8-naphthalimide) bearing the DNA-intercalating moiety NP was designed. The structure of PtNP was fully characterized by single-crystal X-ray crystallography, NMR, and HRMS. PtNP is superior to cisplatin in both in vitro and in vivo anticancer activities with low systemic toxicity. The interaction of PtNP with CT-DNA demonstrated that PtNP could effectively bind to DNA through both covalent and noncovalent double binding modes. In addition to causing significant damage to nDNA and remarkable inhibition to DNA damage repair, PtNP also distributed in mitochondria, inducing mtDNA damage and affecting the downstream transcriptional level of mitochondrion-encoded genes. In addition, PtNP disturbed the physiological processes of mitochondria by reducing the mitochondrial membrane potential and promoting the generation of reactive oxygen species. Mechanistic studies demonstrate that PtNP induced apoptosis via mitochondrial pathways by upregulating Bax and Puma and downregulating Bcl-2 proteins, leading to the release of cytochrome c and activation of caspase-3 and caspase-9. As a dual-DNA-damage agent, PtNP is able to improve the anticancer activity by damaging both nuclear and mitochondrial DNA, thus providing a new anticancer mechanism of action for the naphthalimide monofunctional platinum(II) complexes.


Assuntos
Antineoplásicos/farmacologia , Dano ao DNA/efeitos dos fármacos , Substâncias Intercalantes/farmacologia , Compostos Organoplatínicos/farmacologia , Animais , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Cristalografia por Raios X , Humanos , Substâncias Intercalantes/química , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos Endogâmicos ICR , Modelos Moleculares , Naftalimidas/química , Naftalimidas/farmacologia , Neoplasias/tratamento farmacológico , Neoplasias/genética , Compostos Organoplatínicos/química , Piridinas/química , Piridinas/farmacologia
20.
Mater Sci Eng C Mater Biol Appl ; 105: 110079, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31546406

RESUMO

In the present paper, we synthesized and characterized four N-donor polypyridyl copper(II) complexes (C1-C4); [Cu(mono-CN-PIP)2]2+ (C1), [Cu(tri-OMe-PIP)2]2+ (C2), [Cu(di-CF3-PIP)2]2+ (C3) and [Cu(DPPZ)2]2+ (C4). The (Calf-Thymus) CT-DNA binding studies depicted that the complexes could interact with DNA via intercalative mode. All the complexes, particularly C3 and C4 attenuated the proliferation as well as migration of various cancer cells, indicating their anti-cancer and anti-metastatic activity. Additionally, chick embryo angiogenesis (CEA) assay exhibited the inhibition of vascular sprouting in presence of C3 and C4, suggesting their potential in inhibiting the blood vessel growth. Mechanistic studies revealed that the complexes induced the excessive production of cellular reactive oxygen species (ROS) leading to apoptosis through up regulation of p53 and downregulation of Bcl-xL, which might be the plausible mechanisms underlying their anti-cancer properties. To understand the feasibility of practical application of anti-cancer copper complexes C3 and C4, in vivo sub-chronic toxicity study (4 weeks) was performed in C57BL6 mice and the results exhibited almost non-toxic effects induced by these complexes in terms of haematology and serum biochemical analyses, suggesting their biocompatible nature. The current study provides the basis for future advancement of other novel biocompatible metal complexes that could be employed for the therapy of different cancers.


Assuntos
Complexos de Coordenação , Cobre , Substâncias Intercalantes , Melanoma Experimental , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacocinética , Linhagem Celular Tumoral , Embrião de Galinha , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Cobre/química , Cobre/farmacologia , Humanos , Substâncias Intercalantes/síntese química , Substâncias Intercalantes/química , Substâncias Intercalantes/farmacologia , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Proteína Supressora de Tumor p53/metabolismo , Proteína bcl-X/metabolismo
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