RESUMEN
Conjugate the fluorescent dye 4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indatsen-8-propionic acid (BODIPY) and N2,N4,N6-trimethylmelamine was obtained. It was shown that this compound in the presence of formaldehyde generates covalent cross-links of DNA strands in vitro.
Asunto(s)
Antineoplásicos/síntesis química , Compuestos de Boro/síntesis química , Reactivos de Enlaces Cruzados/síntesis química , Colorantes Fluorescentes/síntesis química , Triazinas/síntesis química , Animales , Antineoplásicos/uso terapéutico , ADN/química , Humanos , Neoplasias/tratamiento farmacológico , Espectrometría de FluorescenciaRESUMEN
An efficient synthetic method for monomer ribonucleotide synthons containing 2'-O-methoxymethyl and 2'O-(p-nitrobenzyloxy)methyl groups used for oligonucleotide phosphotriester method with O-nucleophilic intramolecular catalysis at the stage of formation of internucleotide bond is developed. It is shown that synthons containing protecting 2'-O-(p-nitrobenzyloxy)methyl group may be used for automatic synthesis of phosphotriester oligoribonucleotides with high yields and synthons containing methoxymethyl group--to get 2'-O-modified oligonucleotides.
Asunto(s)
Éteres Metílicos/síntesis química , Nitrobencenos/síntesis química , Oligorribonucleótidos/síntesis química , Hidrolasas de Triéster Fosfórico/metabolismo , Animales , Catálisis , Humanos , Biología Molecular/métodosRESUMEN
A general scheme for the synthesis of ribonucleotide monomers containing alkoxymethyl group in 2'-O-position for the solid-phase phosphotriester oligonucleotide synthesis using O-nucleophilic intramolecular catalysis has been developed. In particular, the monomers containing 2'-O-modifying 2-azidoethoxymethyl, propargyloxymethyl, or 3,4-cyclocarbonatebutoxymethyl groups has been prepared.
Asunto(s)
Oligorribonucleótidos/síntesis química , Ribonucleótidos/química , Técnicas de Síntesis en Fase Sólida , Aldehídos/química , Alquinos/química , Azidas/química , Catálisis , Estructura MolecularRESUMEN
An effective modification of phosphotriester method for automatic synthesis of DNA and RNA fragments using O-nucleophilic intramolecular catalysis and 2-(azidometil)benzoyl group to protect amino groups of heterocyclic bases of nucleotides is described.
Asunto(s)
Azidas/química , Oligonucleótidos/síntesis química , Compuestos Organofosforados/química , Catálisis , Electroforesis en Gel de Poliacrilamida , Estructura Molecular , Oligonucleótidos/químicaRESUMEN
Published data on the main types of reagents capable of introducing covalent interstrand cross links into nucleic acids (NA) are summarized in the present review. The reactivity of cross-linking agents, their preferred binding sites, and methods of determining the cross-link localization in a duplex are discussed. Cell response to DNA cross linking, namely, the blocking of replication and transcription, the initiation of reparation processes, and apoptotic death of the cell, are analyzed, as well as the use of cross-linking reagents in therapy and molecular biology.
Asunto(s)
Reactivos de Enlaces Cruzados , ADN , ARN , Alquilantes/química , Alquilantes/farmacología , Alquilación , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Apoptosis , Reactivos de Enlaces Cruzados/química , Reactivos de Enlaces Cruzados/metabolismo , Reactivos de Enlaces Cruzados/farmacología , ADN/química , ADN/metabolismo , Humanos , Ácidos Nucleicos Heterodúplex/química , Ácidos Nucleicos Heterodúplex/metabolismo , ARN/química , ARN/metabolismoRESUMEN
This review includes literature data on main types of reagents inducing covalent cross-links of nucleic acids. Reactivity of cross-linking agents, preferable sites for their binding, and methods for determination of location of cross-links in duplex are discussed. Biological responses of cells to cross-linking in nucleic acids, i.e. replication and transcription blocking, onset of repair processes, and apoptotic cell death are considered, as well as application of cross-linking reagents as medicinal drugs for solving molecular-biological problems.
Asunto(s)
Reactivos de Enlaces Cruzados , Ácidos Nucleicos , Reactivos de Enlaces Cruzados/química , Reactivos de Enlaces Cruzados/aislamiento & purificación , Reactivos de Enlaces Cruzados/metabolismo , Humanos , Estructura Molecular , Ácidos Nucleicos/química , Ácidos Nucleicos/aislamiento & purificación , Ácidos Nucleicos/metabolismo , EstereoisomerismoRESUMEN
In order to improve physicochemical and biological properties of natural oligonucleotides in particular increasing their affinity for nucleic acids, the selectivity of action and biological sustainability, several types of DNA mimics were designed. The survey collected data on the synthesis and properties of the DNA mimics - peptide-nucleic acids analogues, which are derivatives of pyrrolidine and hydroxyproline. We examine some physicochemical and biological properties of negatively charged mimics of this type, containing phosphonate residues, and possessing a high affinity for DNA and RNA, selective binding with nucleic acids and stability in various biological systems. Examples of the use of these mimics as tools for molecular biological research, particularly in functional genomics are given. The prospects for their use in diagnostics and medicine are discussed.
Asunto(s)
Materiales Biomiméticos , ADN , Hidroxiprolina/química , Ácidos Nucleicos de Péptidos , Pirrolidinas/química , Animales , Materiales Biomiméticos/síntesis química , Materiales Biomiméticos/química , Biomimética/métodos , Humanos , Ácidos Nucleicos de Péptidos/síntesis química , Ácidos Nucleicos de Péptidos/químicaRESUMEN
A rapid and effective method of an automatic oligoribonucleotide synthesis alternative to the phosphoroamidite one was developed based on the phosphotriester approach of internucleotide bond formation under intramolecular O-nucleophilic catalysis and the use of an azidomethyl group for protection of a nucleotide 2'-hydroxyl function.
Asunto(s)
Oligorribonucleótidos/química , Oligorribonucleótidos/síntesis química , CatálisisRESUMEN
The azidomethyl and 2-(azidomethyl)benzoyl as 2'-OH protecting groups are reported for preparation of oligoribonucleotides by the phosphotriester solid-phase method using O-nucleophilic intramolecular catalysis. The procedures for the synthesis of the corresponding monomer synthons were developed and the usefulness of the application of 2'-O-azidomethyl and 2'-O-2-(azidomethyl)benzoyl groups was examined in the synthesis of different RNA fragments with a chain length of 15-22 nucleotides. The azidomethyl group was found to be more preferable for effective synthesis of oligoribonucleotides. Hybridization properties of RNAs toward their complementary oligonucleotides were examined before and after the removal of 2'-O-azidomethyl groups.
Asunto(s)
Azidas/química , Oligorribonucleótidos/síntesis química , ARN/síntesis química , Secuencia de Bases , Ensayo de Cambio de Movilidad Electroforética , Datos de Secuencia Molecular , Estructura Molecular , Oligorribonucleótidos/química , Compuestos Organofosforados/química , ARN/químicaRESUMEN
Morpholino phosphorodiamidate (MO) DNA mimics display excellent water solubility and hybridization properties toward DNA and RNA, and have been utilized in the model vertebrate zebrafish (Danio rerio) for genome-wide, sequence-based, reverse genetic screens during embryonic development. Peptide nucleic acids (PNAs) exhibit excellent mismatch discrimination, nuclease resistance, and protease resistance, but low solubility. Negatively charged DNA mimics composed of alternating residues of trans-4-hydroxy-L-proline peptide nucleic acid monomers and phosphono peptide nucleic acid monomers (HypNA-pPNA) combine all of the positive features of both MOs and PNAs. Thus, we evaluated PNA oligomers and HypNA-pPNA oligomers as an alternative to MOs for oligonucleotide inhibition of gene expression in zebrafish embryos. We observed that HypNA-pPNA 18-mers displayed comparable potency to MO 25-mers as knockdown agents against chordin, notail and uroD, with greater mismatch stringency. Furthermore, we observed that a specific HypNA-pPNA 18-mer elicited the dharma (bozozok)(-/-) phenotype in zebrafish embryos, which MO 25-mers do not. These observations validate HypNA-pPNAs as an alternative to MO oligomers for reverse genetic studies. The stronger hybridization and greater specificity of HypNA-pPNAs enable knockdown of mRNAs unaffected by MO oligomers.
Asunto(s)
Embrión no Mamífero/efectos de los fármacos , Hidroxiprolina/química , Oligorribonucleótidos Antisentido/farmacología , Organofosfonatos/química , Ácidos Nucleicos de Péptidos/química , ARN Mensajero/antagonistas & inhibidores , Pez Cebra/embriología , Animales , Secuencia de Bases , Embrión no Mamífero/metabolismo , Imitación Molecular , Oligorribonucleótidos Antisentido/química , ARN Mensajero/biosíntesisRESUMEN
An effective procedure for specific determination of the cap structure at the 5'-terminus of mRNA and for isolation of the corresponding full-length cDNA has been developed. The procedure involves covalent attachment of an oligonucleotide template extender to the 5'-cap structure of mRNA followed by RT-PCR using M-MLV SuperScript II reverse transcriptase. In the course of reverse transcription, the enzyme 'jumps over' the cap structure and includes the sequence complementary to the oligonucleotide template extender into the 3'-end of the first cDNA strand. The cap-jumping method was successfully tested using some mammalian cellular mRNAs, genomic RNAs of tobacco mosaic virus (TMV) U1 and the recently isolated crucifer-infecting tobamovirus. Moreover, cDNA products corresponding to the genomic tobamovirus RNA were obtained from total RNA extracted from tobacco plants infected by crucifer-infecting tobamovirus or tobacco mosaic virus. Using the cap-jumping method, we have shown for the first time that genomic crucifer-infecting tobamovirus (crTMV) RNA contains a 5'-cap structure. This improved method can be recommended for the construction of full-length and 5'-end enriched cDNA libraries, identification of capped RNAs and determination of their 5'-terminal sequences.
Asunto(s)
Caperuzas de ARN/genética , ARN Mensajero/genética , Regiones no Traducidas 5'/química , Regiones no Traducidas 5'/genética , Animales , Secuencia de Bases , ADN Complementario/genética , Globinas/genética , Oligonucleótidos/química , Oligonucleótidos/genética , Caperuzas de ARN/química , ARN Mensajero/química , ARN Mensajero/metabolismo , ARN Viral/química , ARN Viral/genética , ARN Viral/metabolismo , ADN Polimerasa Dirigida por ARN/metabolismo , Conejos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tobamovirus/genéticaRESUMEN
Effects of low activity of test-subjects during antiorthostatic hypokinesia on the intragroup structure as well as the semantic composition and the directionality of estimation criteria selected by test-subjects to describe the system of attitudes to oneself and others, and their impact on individual status in the social structure of the group have been studied. It was shown that maintaining of a definite level of individual activity of members is important for the group structure and successful adaptation of test-subjects to the unfavourable factors of hypokinesia. The degree of the ego image integrity and dominance of criteria that infer empathy and positive feelings within the system of individual appraisements determine the social status of a group member. The psychologic signs of a "apet" (psychologically close) and a "outsider" (psychologically strange) members of the group have been revealed.
Asunto(s)
Adaptación Psicológica , Hipocinesia/psicología , Relaciones Interpersonales , Vuelo Espacial , Adulto , Codependencia Psicológica , Humanos , Masculino , Factores de Tiempo , IngravidezRESUMEN
Two types of oligonucleotide mimics relative to peptide nucleic acids (PNAs) were tested as probes in nucleic acid hybridisation assays based on polyacrylamide technology. One type of mimic oligomers represented a chimera constructed of PNA and phosphono-PNA (pPNA) monomers, and the other one contained pPNA residues alternating with PNA-like monomers on the base of trans -4-hydroxy-L-proline (HypNA). A chemistry providing efficient and specific covalent attachment of these DNA mimics to acrylamide polymers using a convenient approach based on the co-polymerisation of acrylamide and some reactive acrylic acid derivatives with oligomers bearing 5'- or 3'-terminal acrylamide groups has been developed. A comparative study of polyacrylamide conjugates with oligonucleotides and mimic oligomers demonstrated the suitability and high potential of PNA-pPNA and HypNA-pPNA chimeras as sequence-specific probes in capture and detection of target nucleic acid fragments to serve current forms of DNA arrays.
Asunto(s)
Resinas Acrílicas/síntesis química , Imitación Molecular , Oligonucleótidos/síntesis química , Ácidos Nucleicos de Péptidos/química , Acrilamida/química , Resinas Acrílicas/química , ADN/análisis , ADN/aislamiento & purificación , Técnicas y Procedimientos Diagnósticos , Hibridación de Ácido Nucleico , Sondas de Oligonucleótidos , Oligonucleótidos/química , Ácidos Nucleicos de Péptidos/síntesis químicaRESUMEN
Approaches to preparing acrylamide and polyacrylamide conjugates with oligonucleotides and some peptide nucleic acid-related DNA mimics are considered. Their physicochemical properties and application to the nucleic acid analysis are discussed.
Asunto(s)
Resinas Acrílicas/química , ADN/química , Imitación Molecular , Secuencia de Bases , Estructura Molecular , Ácidos Nucleicos de Péptidos/químicaRESUMEN
The objective of this paper is to describe psychological phenomena which has developed in the course of inter- and intragroup interaction in space. Several objective reasons are stressed which comprise the basis of the formation of the majority of the psychological phenomena. The authors also discuss prediction of these phenomena, namely, can they occur on the International Space Station (ISS) and what countermeasures will prevent them. In the analysis, the authors rely on the practice of prolonged spaceflight (PSF) psychological support in Russia; preliminary results of several flights within the Mir/NASA project; the personal experience of cosmonaut Valery V. Polyakov; and data gained during prolonged studies simulating PSF conditions (isolation, microgravity, etc.).
Asunto(s)
Adaptación Psicológica , Astronautas/psicología , Procesos de Grupo , Relaciones Interpersonales , Vuelo Espacial , Astronautas/educación , Diversidad Cultural , Humanos , Federación de Rusia , Aislamiento Social , Estados Unidos , IngravidezRESUMEN
The synthesis of a series of DNA mimics--peptide nucleic acids, phosphonate analogues of peptide nucleic acids, and their hybrids--is described. The preparative synthesis of the corresponding monomers and the solid phase automated synthesis of oligomers-mimics are developed. Modified phosphonate analogues of peptide nucleic acids, in particular chiral derivatives and those with additional hydroxyl groups in the side chains of the backbone as well as pyrene derivatives of peptide nucleic acids and their phosphonate analogues, are prepared. The ability of the resulting oligomers specifically to hybridize to DNA and RNA complementary chains is studied. It is shown that phosphonate analogues of peptide nucleic acids and their hybrids with peptide nucleic acids can form complexes with the DNA and RNA complementary strands, the stability of the complexes increasing in parallel with the increase in the number of peptide nucleic acid residues in the chain of the mimic. This property, along with good water solubility, provides the precondition for further evaluation of these compounds as antisense and antigene agents.
Asunto(s)
ADN/química , Organofosfonatos/química , Ácidos Nucleicos de Péptidos/síntesis química , Cromatografía Líquida de Alta Presión , Imitación Molecular , Hibridación de Ácido Nucleico , Ácidos Nucleicos de Péptidos/química , ARN/químicaRESUMEN
Two convenient, practical routes to the synthesis of non-nucleotide bridged cyclic oligonucleotides have been developed. The first procedure included circularization of oligonucleotides by template-directed ligation on solid phase, while the second procedure involved preparation of a circular oligomer by non-template chemical ligation of a linear precursor in solution. Using these approaches, a series of single- and double-stranded cyclic oligonucleotides with non-nucleotide bridges has been synthesized.
Asunto(s)
ADN Circular/síntesis química , Oligonucleótidos/síntesis química , Fármacos Anti-VIH , Línea Celular , Humanos , Inhibidores de Integrasa/síntesis química , Conformación Molecular , Estructura Molecular , Conformación de Ácido Nucleico , Desnaturalización de Ácido Nucleico , Hibridación de Ácido Nucleico , Glicoles de Propileno/síntesis química , Transcripción Genética/efectos de los fármacos , Transfección/genéticaRESUMEN
BACKGROUND: The interactions of individuals participating in space simulation studies can give us valuable information about ways that space crew members relate with one another. A promising method of investigating such interactions is based on the Kelly repertory grid technique. HYPOTHESES: We predicted that in a space simulation crew: 1) one's personal self-concept would become closer to concepts about other crew members in a cohesive crew, and vice-versa, and 2) the similarity of one's ego images in the past, present and future influences that person's position in the group. METHOD: Crews from two isolation studies lasting 135 and 90 days at the Institute for Biomedical Problems in Moscow were evaluated using a modification of the Kelly repertory grid technique. Members assessed each other on a monthly basis and during off-nominal simulated docking periods. RESULTS: During both isolations, crew members were unsuccessful in making their personal self-concepts move closer to their concepts about fellow crew members. Crew disintegration resulted, with one member becoming an outsider whose personality was characterized by a disintegrated ego-image and an infantile type of the present ego. CONCLUSIONS: In small isolated groups, failure to make personal self-concepts become more like concepts about other crew members can lead to group disintegration and tension and can result in the appearance of an outsider who has a disintegrated ego-image and an infantile present ego.
Asunto(s)
Autoimagen , Aislamiento Social , Vuelo Espacial , Adulto , Ego , Humanos , Relaciones Interpersonales , Masculino , Persona de Mediana EdadRESUMEN
In an attempt to improve physico-chemical and biological properties of peptide nucleic acids (PNAs), particularly water solubility and cellular uptake, the synthesis of chimeric oligomers consisted of PNA and phosphono-PNA analogues (pPNAs) bearing the four natural nucleobases has been accomplished. To produce these chimeras, pPNA monomers of two types containing N-(2-hydroxyethyl)phosphonoglycine, or N-(2-aminoethyl)phosphonoglycine backbone, were used in conjunction with PNA monomers representing derivatives of N-(2-aminoethyl)glycine, or N-(2-hydroxyethyl)glycine. The oligomers obtained were composed of either PNA and pPNA stretches or alternating PNA and pPNA monomers. The examination of hybridization properties of PNA-pPNA chimeras to DNA and RNA complementary strands in comparison with pure PNAs, and pPNAs as well as DNA-pPNA hybrids and DNA fragments confirmed that these chimeras form stable complexes with complementary DNA and RNA fragments. They were found to be resistant to degradation by nucleases. All these properties together with good solubility in water make PNA-pPNA hybrids promising for further evaluation as potential therapeutic agents.
Asunto(s)
Ácidos Nucleicos/síntesis química , Oligonucleótidos/síntesis química , Péptidos/síntesis química , Fenómenos Químicos , Química Física , ADN Complementario/química , Dimerización , Estabilidad de Medicamentos , Glicina/análogos & derivados , Glicina/química , Calor , Hibridación de Ácido Nucleico , Ácidos Nucleicos/química , Ácidos Nucleicos/metabolismo , Nylons/química , Organofosfonatos/química , Péptidos/química , Péptidos/metabolismo , Fosfodiesterasa I , Hidrolasas Diéster Fosfóricas/metabolismo , ARN Complementario/química , Endonucleasas Específicas del ADN y ARN con un Solo Filamento/metabolismo , Solubilidad , Rayos UltravioletaRESUMEN
Thymidine oligonucleotide analogues with phosphodiester bonds fully substituted by carboxamidomethyl phosphonamide, or glycinamide linkages were synthesized on a solid support, and their hybridization properties toward DNA and RNA targets were determined by Tm analysis.