RESUMEN
Fludarabine (FA) is still considered as a first-line chemotherapy drug for hematological tumors related to B lymphocytes. However, it is worth noting that the non-specific distribution and non-different cytotoxicity of FA may lead to irreversible consequences such as central nervous system damage such as blindness, coma, and even death. Therefore, it is very important to develop a system to targeting delivery FA. In preliminary studies, it was found that B lymphoma cells would specific highly expressing the sialic acid-binding immunoglobulin-like lectin 2 (known as CD22). Inspired by the specific recognition of sialic acid residues and CD22, we have developed a supramolecular prodrug based on polysialic acid, an endogenous biomacromolecule, achieving targeted-therapy of B-cell non-Hodgkin's lymphoma (B-NHL). Specifically, the prepared hydrophobic reactive oxygen species-responsive FA dimeric prodrug (F2A) interacts with the TPSA, which polysialic acid were modified by the thymidine derivatives, through non-covalent intermolecular interactions similar to "Watson-Crick" base pairing, resulting in the formation of nanoscale supramolecular prodrug (F@TPSA). Cell experiments have confirmed that F@TPSA can be endocytosed by CD22+ B lymphoma cells including Raji and Ramos cells, and there is a significant difference of endocytosis in other leukocytes. Furthermore, in B-NHL mouse model, compared with FA, F@TPSA is determined to have a stronger tumor targeting and inhibitory effect. More importantly, the distribution of F@TPSA in vivo tends to be enriched in lymphoma tissue rather than nonspecific, thus reducing the leukopenia of FA. The targeted delivery system based on PSA provides a new prodrug modification strategy for targeted treatment of B-NHL.
Asunto(s)
Linfoma de Células B , Profármacos , Profármacos/química , Profármacos/farmacología , Animales , Ratones , Humanos , Línea Celular Tumoral , Linfoma de Células B/tratamiento farmacológico , Ácidos Siálicos/química , Ácidos Siálicos/farmacología , Lectina 2 Similar a Ig de Unión al Ácido Siálico , Antineoplásicos/farmacología , Antineoplásicos/química , Nanopartículas/química , Medicina de Precisión/métodos , Sistemas de Liberación de Medicamentos/métodos , Ratones Endogámicos BALB C , Especies Reactivas de Oxígeno/metabolismo , Linfoma no Hodgkin/tratamiento farmacológicoRESUMEN
Base pairing in RNA are significantly rich and versatile due to the potential non-canonical base pairing amongst nucleotides. Not only that, one base in RNA can pair with more than one bases simultaneously. This opens up a new dimension of research to detect such types of base-base pair networks in RNA and to analyze them. Even if a base do not form a pair, it may have significant extent of [Formula: see text]-[Formula: see text] stacking overlap that can stabilize the structures. In this work, we report a software tool, called BPNet, that accepts a mmCIF or PDB file and computes the base-pair/[Formula: see text]-[Formula: see text] contact network components using graph formalism. The software can run on Linux platform in both serial and parallel modes. It generates several information in suitable file formats for visualization of the networks. This paper describes the BPNet software and also presents some interesting results obtained by analyzing several RNA structures by the software to show its effectiveness.
Asunto(s)
Biología Computacional , ARN , Emparejamiento Base , Enlace de Hidrógeno , Conformación de Ácido Nucleico , ARN/químicaRESUMEN
Historians and biologists identify the debate between mechanists and vitalists over the nature of life itself with the arguments of Driesch, Loeb, and other prominent voices. But what if the conversation was broader and the consequences deeper for the field? Following the suspicions of Joseph Needham in the 1930s and Francis Crick in the 1960s, we deployed tools of the digital humanities to an old problem in the history of biology. We analyzed over 31,000 peer-reviewed scientific papers and learned that bioexceptionalism participated in a robust discursive landscape throughout subfields of the life sciences, occupied even by otherwise unknown biologists.
Asunto(s)
Disciplinas de las Ciencias Biológicas , Biología , Comunicación , Historia del Siglo XX , Humanidades , VitalismoRESUMEN
Energetically unfavorable Watson-Crick (WC)-like tautomeric forms of nucleobases are known to introduce spontaneous mutations, and contribute to replication, transcription, and translation errors. Recent NMR relaxation dispersion techniques were able to show that wobble (w) Gâ¢U mispair exists in equilibrium with the short-lived, low-population WC-like enolic tautomers. Presently, we have investigated the wGâ¢U â WC-like enolic reaction pathway using various theoretical methods: quantum mechanics (QM), molecular dynamics (MD), and combined quantum mechanics/molecular mechanics (QM/MM). The previous studies on QM gas phase calculations were inconsistent with experimental data. We have also explored the environmental effects on the reaction energies by adding explicit water. While the QM-profile clearly becomes endoergic in the presence of water, the QM/MM-profile remains consistently endoergic in the presence and absence of water. Hence, by including microsolvation and QM/MM calculations, the experimental data can be explained. For the Gâ¢Uenolâ Genolâ¢U pathway, the latter appears to be energetically more favorable throughout all computational models. This study can be considered as a benchmark of various computational models of wGâ¢U to WC-like tautomerization pathways with and without the environmental effects, and may contribute on further studies of other mispairs as well.
Asunto(s)
Guanina/metabolismo , ARN/genética , Uracilo/metabolismo , Disparidad de Par Base/genética , Emparejamiento Base/genética , Simulación por Computador , Modelos Moleculares , Simulación de Dinámica Molecular , Mutación Puntual/genética , Teoría CuánticaRESUMEN
DNA duplexes comprising 6-(1H-pyrazol-1-yl)-9H-purine (6PP), 1-deaza-6PP (1D 6PP), 7-deaza-6PP (7D 6PP) and 1,7-dideaza-6PP (1,7D 6PP) 2'-deoxyribonucleosides, respectively, were investigated towards their ability to form metal-mediated base pairs in the presence of AgI . In 6PP and 7D 6PP, the AgI ion can coordinate to the nucleobase via the endocyclic N1 nitrogen atom, that is, via the Watson-Crick edge. In contrast, this nitrogen atom is not available in 1D 6PP and 1,7D 6PP, so that in 1D 6PP an AgI coordination is only possible via the Hoogsteen edge (N7). Reference duplexes with either adenine:adenine mispairs or canonical adenine:thymine base pairs were used to investigate the impact of the pyrazolyl moiety on the AgI -binding properties. To determine the thermal and structural duplex stabilities in the absence or presence of AgI , all duplexes were examined by UV and circular dichroism spectroscopic studies. These investigations shed light on the question of whether N1- or N7-coordination is preferred in purine-based metal-mediated base pairs.
Asunto(s)
Complejos de Coordinación , ADN , Purinas , Pirazoles , Plata , Emparejamiento Base , Complejos de Coordinación/química , ADN/química , Purinas/química , Pirazoles/química , Plata/químicaRESUMEN
A theoretical study of the effect of the diprotonation on the nucleic acid bases (A : U, A : T and G : C) in Watson-Crick conformation has been carried out by means of DFT computational methods in vacuum. In addition, the corresponding neutral and monoprotonated binary complexes have been considered. Most of the diprotonated species studied are stable, even though the binding energy is positive due to the overall repulsive electrostatic term. Local electrostatic attractive forces in the regions of hydrogen bonds (HBs) are responsible for equilibrium geometries, as shown by the electric field lines connecting the electrophilic and nucleophilic sites involved in the HB interactions. Secondary electrostatic effects also affect the assembling of the nucleic acid complexes in either neutral or cationic form. In particular, the electric field lines flowing from electrophilic sites in one base to nucleophilic sites in the other reinforce the linking between them. Hence, when the nucleophilic site concerns the free lone pair of the heteroatom involved in the HB interaction as acceptor, the HB distance shortens. However, if the free lone pair of the HB acceptor interacts with an electrophilic site in the same molecule, the HB distance elongates, weakening the HB interaction. The topological analysis of the electron density distribution in HB regions indicates that neutral, monoprotonated and diprotonated complexes show no differences in the nature of their HB's.
Asunto(s)
Modelos Moleculares , Ácidos Nucleicos/química , Emparejamiento Base , Cationes , Enlace de Hidrógeno , Conformación de Ácido Nucleico , Teoría Cuántica , Electricidad Estática , TermodinámicaRESUMEN
The emergence of unnatural DNA bases provides opportunities to demystify the mechanisms by which DNA polymerases faithfully decode chemical information on the template. It was previously shown that two unnatural cytosine bases (termed "M-fC" and "I-fC"), which are chemical labeling adducts of the epigenetic base 5-formylcytosine, can induce C-to-T transition during DNA amplification. However, how DNA polymerases recognize such unnatural cytosine bases remains enigmatic. Herein, crystal structures of unnatural cytosine bases pairing to dA/dG in the KlenTaq polymerase-host-guest complex system and pairing to dATP in the KlenTaq polymerase active site were determined. Both M-fC and I-fC base pair with dA/dATP, but not with dG, in a Watson-Crick geometry. This study reveals that the formation of the Watson-Crick geometry, which may be enabled by the A-rule, is important for the recognition of unnatural cytosines.
Asunto(s)
Citosina/química , ADN Polimerasa Dirigida por ADN/química , ADN/química , Timina/química , Humanos , Estructura MolecularRESUMEN
DNA sequences comprising noncanonical 7-deazaguanine (7C G) and canonical cytosine (C) are capable of forming Watson-Crick base pairs via hydrogen bonds as well as silver(I)-mediated base pairs by coordination to central silver(I) ions. Duplexes I and II containing 7C G and C have been synthesized and characterized. The incorporation of silver(I) ions into these duplexes has been studied by means of temperature-dependent UV spectroscopy, circular dichroism, and DFT calculations. The results suggest the formation of DNA molecules comprising contiguous metallated 7C G-AgI -C Watson-Crick base pairs that preserve the original B-type conformation. Furthermore, additional studies performed on duplex III indicated that, in the presence of AgI ions, 7C G-C and 7C A-T Watson-Crick base pairs (7C A, 7-deazadenine; T, thymine) can be converted to metallated 7C G-AgI -C and 7C A-AgI -T base pairs inside the same DNA molecule whilst maintaining its initial double helix conformation. These findings are very important for the development of customized silver-DNA nanostructures based on a Watson-Crick complementarity pattern.
Asunto(s)
Citosina/química , ADN/química , Guanina/análogos & derivados , Plata/química , Emparejamiento Base , Secuencia de Bases , Guanina/químicaRESUMEN
We synthesized a novel linker (1) with biotin, alkyne and amino groups for the identification of target proteins using a small molecule that contains an azide group (azide probe). The alkyne in the linker bound the azide probe via an azide-alkyne Huisgen cycloaddition. A protein cross-linker effectively bound the conjugate of the linker and an azide probe with a target protein. The covalently bound complex was detected by western blotting. Linker 1 was applied to a model system using an abscisic acid receptor, RCAR/PYR/PYL (PYL). Cross-linked complexes of linker 1, the azide probes and the target proteins were successfully visualized by western blotting. This method of target protein identification was more effective than a previously developed method that uses a second linker with biotin, alkyne, and benzophenone (linker 2) that acts to photo-crosslink target proteins. The system developed in this study is a method for identifying the target proteins of small bioactive molecules and is different from photo-affinity labelling.
Asunto(s)
Alquinos/química , Proteínas de Arabidopsis/química , Biotina/análogos & derivados , Biotina/química , Sondas Moleculares/química , Alquinos/síntesis química , Arabidopsis/química , Azidas/síntesis química , Azidas/química , Biotina/síntesis química , Western Blotting , Química Clic , Reactivos de Enlaces Cruzados/síntesis química , Reactivos de Enlaces Cruzados/química , Reacción de Cicloadición , Escherichia coli/química , Proteínas de Escherichia coli/química , Oxidorreductasas Intramoleculares/química , Luminiscencia , Lisina/análogos & derivados , Lisina/síntesis química , Lisina/química , Sondas Moleculares/síntesis químicaRESUMEN
Comprehensive analyses of structural features of non-canonical base pairs within a nucleic acid double helix are limited by the availability of a small number of three dimensional structures. Therefore, a procedure for model building of double helices containing any given nucleotide sequence and base pairing information, either canonical or non-canonical, is seriously needed. Here we describe a program RNAHelix, which is an updated version of our widely used software, NUCGEN. The program can regenerate duplexes using the dinucleotide step and base pair orientation parameters for a given double helical DNA or RNA sequence with defined Watson-Crick or non-Watson-Crick base pairs. The original structure and the corresponding regenerated structure of double helices were found to be very close, as indicated by the small RMSD values between positions of the corresponding atoms. Structures of several usual and unusual double helices have been regenerated and compared with their original structures in terms of base pair RMSD, torsion angles and electrostatic potentials and very high agreements have been noted. RNAHelix can also be used to generate a structure with a sequence completely different from an experimentally determined one or to introduce single to multiple mutation, but with the same set of parameters and hence can also be an important tool in homology modeling and study of mutation induced structural changes.
Asunto(s)
Modelos Moleculares , Ácidos Nucleicos/química , Emparejamiento Base , Conformación de Ácido Nucleico , Electricidad EstáticaRESUMEN
RNA 3D motifs occupy places in structured RNA molecules that correspond to the hairpin, internal and multi-helix junction "loops" of their secondary structure representations. As many as 40% of the nucleotides of an RNA molecule can belong to these structural elements, which are distinct from the regular double helical regions formed by contiguous AU, GC, and GU Watson-Crick basepairs. With the large number of atomic- or near atomic-resolution 3D structures appearing in a steady stream in the PDB/NDB structure databases, the automated identification, extraction, comparison, clustering and visualization of these structural elements presents an opportunity to enhance RNA science. Three broad applications are: (1) identification of modular, autonomous structural units for RNA nanotechnology, nanobiology and synthetic biology applications; (2) bioinformatic analysis to improve RNA 3D structure prediction from sequence; and (3) creation of searchable databases for exploring the binding specificities, structural flexibility, and dynamics of these RNA elements. In this contribution, we review methods developed for computational extraction of hairpin and internal loop motifs from a non-redundant set of high-quality RNA 3D structures. We provide a statistical summary of the extracted hairpin and internal loop motifs in the most recent version of the RNA 3D Motif Atlas. We also explore the reliability and accuracy of the extraction process by examining its performance in clustering recurrent motifs from homologous ribosomal RNA (rRNA) structures. We conclude with a summary of remaining challenges, especially with regard to extraction of multi-helix junction motifs.
Asunto(s)
ARN/química , Animales , Emparejamiento Base , Simulación por Computador , Bases de Datos de Ácidos Nucleicos , Humanos , Modelos Moleculares , Conformación de Ácido Nucleico , Motivos de Nucleótidos , Programas InformáticosRESUMEN
Protein synthesis must rapidly and repeatedly discriminate between a single correct and many incorrect aminoacyl-tRNAs. We have attempted to measure the frequencies of all possible missense errors by tRNA , tRNA and tRNA . The most frequent errors involve three types of mismatched nucleotide pairs, Uâ¢U, Uâ¢C, or Uâ¢G, all of which can form a noncanonical base pair with geometry similar to that of the canonical Uâ¢A or Câ¢G Watson-Crick pairs. Our system is sensitive enough to measure errors at other potential mismatches that occur at frequencies as low as 1 in 500,000 codons. The ribosome appears to discriminate this efficiently against any pair with non-Watson-Crick geometry. This extreme accuracy may be necessary to allow discrimination against the errors involving near Watson-Crick pairing.
Asunto(s)
Disparidad de Par Base/fisiología , Mutación Missense , Biosíntesis de Proteínas/fisiología , Ribosomas/fisiología , Sustitución de Aminoácidos , Emparejamiento Base/fisiología , Escherichia coli/genética , Escherichia coli/metabolismo , Mutagénesis/fisiología , Mutación Missense/fisiología , Conformación de Ácido Nucleico , Organismos Modificados Genéticamente , ARN de Transferencia de Aspártico/metabolismo , ARN de Transferencia de Ácido Glutámico/metabolismo , ARN de Transferencia de Tirosina/metabolismo , beta-Galactosidasa/química , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
Stacking interaction between the aromatic heterocyclic bases plays an important role in the double helical structures of nucleic acids. Considering the base as rigid body, there are total of 18 degrees of freedom of a dinucleotide step. Some of these parameters show sequence preferences, indicating that the detailed atomic interactions are important in the stacking. Large variants of non-canonical base pairs have been seen in the crystallographic structures of RNA. However, their stacking preferences are not thoroughly deciphered yet from experimental results. The current theoretical approaches use either the rigid body degrees of freedom where the atomic information are lost or computationally expensive all atom simulations. We have used a hybrid simulation approach incorporating Monte-Carlo Metropolis sampling in the hyperspace of 18 stacking parameters where the interaction energies using AMBER-parm99bsc0 and CHARMM-36 force-fields were calculated from atomic positions. We have also performed stacking energy calculations for structures from Monte-Carlo ensemble by Dispersion corrected density functional theory. The available experimental data with Watson-Crick base pairs are compared to establish the validity of the method. Stacking interaction involving A:U and G:C base pairs with non-canonical G:U base pairs also were calculated and showed that these structures were also sequence dependent. This approach could be useful to generate multiscale modeling of nucleic acids in terms of coarse-grained parameters where the atomic interactions are preserved. This method would also be useful to predict structure and dynamics of different base pair steps containing non Watson-Crick base pairs, as found often in the non-coding RNA structures. © 2015 Wiley Periodicals, Inc. Biopolymers 105: 212-226, 2016.
Asunto(s)
Emparejamiento Base , Enlace de Hidrógeno , Modelos Moleculares , Conformación de Ácido Nucleico , Teoría CuánticaRESUMEN
The oligonucleotide d(TX)9 , which consists of an octadecamer sequence with alternating non-canonical 7-deazaadenine (X) and canonical thymine (T) as the nucleobases, was synthesized and shown to hybridize into double-stranded DNA through the formation of hydrogen-bonded Watson-Crick base pairs. dsDNA with metal-mediated base pairs was then obtained by selectively replacing W-C hydrogen bonds by coordination bonds to central silver(I) ions. The oligonucleotide I adopts a duplex structure in the absence of Ag(+) ions, and its stability is significantly enhanced in the presence of Ag(+) ions while its double-helix structure is retained. Temperature-dependent UV spectroscopy, circular dichroism spectroscopy, and ESI mass spectrometry were used to confirm the selective formation of the silver(I)-mediated base pairs. This strategy could become useful for preparing stable metallo-DNA-based nanostructures.
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Adenina/análogos & derivados , ADN/química , Plata/química , Timina/química , Adenina/química , Adenina/metabolismo , Emparejamiento Base , Dicroismo Circular , ADN/metabolismo , Enlace de Hidrógeno , Conformación de Ácido Nucleico , Espectrometría de Masa por Ionización de Electrospray , Espectrofotometría Ultravioleta , Temperatura , Timina/metabolismoRESUMEN
Non random segregation of sister chromatids has been implicated to help specify daughter cell fate (the Silent Sister Hypothesis [1]) or to protect the genome of long-lived stem cells (the Immortal Strand Hypothesis [2]). The idea that sister chromatids are non-randomly segregated into specific daughter cells is only marginally supported by data in sporadic and often contradictory studies. As a result, the field has moved forward rather slowly. The advent of being able to directly label and differentiate sister chromatids in vivo using fluorescence in situ hybridization [3] was a significant advance for such studies. However, this approach is limited by the need for large tracks of unidirectional repeats on chromosomes and the reliance on quantitative imaging of fluorescent probes and rigorous statistical analysis to discern between the two competing hypotheses. A novel method called Strand-seq which uses next-generation sequencing to assay sister chromatid inheritance patterns independently for each chromosome [4] offers a comprehensive approach to test for non-random segregation. In addition Strand-seq enables studies on the deposition of chromatin marks in relation to DNA replication. This method is expected to help unify the field by testing previous claims of non-random segregation in an unbiased way in many model systems in vitro and in vivo.
Asunto(s)
Segregación Cromosómica , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Animales , ADN/metabolismo , Humanos , Hibridación Fluorescente in Situ , Modelos Biológicos , Intercambio de Cromátides HermanasRESUMEN
Non-Watson-Crick pairs like the G·U wobble are frequent in RNA duplexes. Their geometric dissimilarity (nonisostericity) with the Watson-Crick base pairs and among themselves imparts structural variations decisive for biological functions. Through a novel circular representation of base pairs, a simple and general metric scheme for quantification of base-pair nonisostericity, in terms of residual twist and radial difference that can also envisage its mechanistic effect, is proposed. The scheme is exemplified by G·U and U·G wobble pairs, and their predicable local effects on helical twist angle are validated by MD simulations. New insights into a possible rationale for contextual occurrence of G·U and other non-WC pairs, as well as the influence of a G·U pair on other non-Watson-Crick pair neighborhood and RNA-protein interactions are obtained from analysis of crystal structure data. A few instances of RNA-protein interactions along the major groove are documented in addition to the well-recognized interaction of the G·U pair along the minor groove. The nonisostericity-mediated influence of wobble pairs for facilitating helical packing through long-range interactions in ribosomal RNAs is also reviewed.
Asunto(s)
Emparejamiento Base , Conformación de Ácido Nucleico , ARN/química , Sitios de Unión , Modelos Moleculares , Simulación de Dinámica Molecular , Proteínas de Unión al ARN/químicaRESUMEN
Emergence of thousands of crystal structures of noncoding RNA molecules indicates its structural and functional diversity. RNA function is based upon a large variety of structural elements which are specifically assembled in the folded molecules. Along with the canonical Watson-Crick base pairs, different orientations of the bases to form hydrogen-bonded non-canonical base pairs have also been observed in the available RNA structures. Frequencies of occurrences of different non-canonical base pairs in RNA indicate their important role to maintain overall structure and functions of RNA. There are several reports on geometry and energetic stabilities of these non-canonical base pairs. However, their stacking geometry and stacking stability with the neighboring base pairs are not well studied. Among the different non-canonical base pairs, the G:U wobble base pair (G:U W:WC) is most frequently observed in the RNA double helices. Using quantum chemical method and available experimental data set we have studied the stacking geometry of G:U W:WC base pair containing dinucleotide sequences in roll-slide parameters hyperspace for different values of twist. This study indicates that the G:U W:WC base pair can stack well with the canonical base pairs giving rise to large interaction energy. The overall preferred stacking geometry in terms of roll, twist and slide for the eleven possible dinucleotide sequences is seen to be quite dependent on their sequences.
Asunto(s)
Emparejamiento Base/fisiología , ARN/química , Enlace de Hidrógeno , Conformación de Ácido NucleicoRESUMEN
Biological and artificial molecules and assemblies capable of supramolecular recognition, especially those with nucleobase pairing, usually rely on autonomous or collective binding to function. Advanced site-specific recognition takes advantage of cooperative spatial effects, as in local folding in protein-DNA binding. Herein, we report a new nucleobase-tagged metal-organic framework (MOF), namely ZnBTCA (BTC=benzene-1,3,5-tricarboxyl, A=adenine), in which the exposed Watson-Crick faces of adenine residues are immobilized periodically on the interior crystalline surface. Systematic control experiments demonstrated the cooperation of the open Watson-Crick sites and spatial effects within the nanopores, and thermodynamic and kinetic studies revealed a hysteretic host-guest interaction attributed to mild chemisorption. We further exploited this behavior for adenine-thymine binding within the constrained pores, and a globally adaptive response of the MOF host was observed.
Asunto(s)
ADN/química , Metales/química , Compuestos Orgánicos/química , Cinética , Espectrofotometría Infrarroja , Espectrometría Raman , TermodinámicaRESUMEN
The structure of Watson-Crick-type adenine-thymine and guanine-cytosine pairs has been studied by hybrid Monte Carlo (HMC) and path integral hybrid Monte Carlo (PIHMC) simulations with the use of semiempirical PM6-DH+ method in the gas phase. We elucidated the nuclear quantum effect and temperature dependency on the hydrogen-bonded moiety of base pairs. It was shown that the contribution of nuclear quantum effect on the hydrogen-bonded structure is significant not only at low temperature 150 K but also at temperature as high as 450 K. The relative position of hydrogen-bonded proton between two heavy atoms and the nuclear quantum nature of the proton are also shown. Furthermore, we have applied principal component analysis to HMC and PIHMC simulations to analyze the nuclear quantum effect on intermolecular motions. We found that the ratio of Buckle mode (lowest vibrational mode from normal mode analysis) decreases due to the nuclear quantum effect, whereas that of Propeller mode (second lowest vibrational mode) increases. In addition, nonplanar structures of base pairs were found to become stable due to the nuclear quantum effect from two-dimensional free energy landscape along Buckle and Propeller modes.