Polarity modified triplex DNA by bromide

Conclusion A py-pu-py triplex DNA containing the polarity modification by replacing the atom H5 of cytosine with Br was examined, mainly for the stability of the formation of the triplex DNA. Our computational results show that the conformation of the strand that contained a replaced H5 by cytosine to Br was destabilized, but the Hoogsteen base pairs were stabilized. Although this approach of modifying the polarity of DNA base has not been tried before, it shows the possibility to improve the stability by the polarity modification.     

Cleavage of pyrimidine dimers in specific DNA sequences by a pyrimidine dimer DNA-glycosylase of M. luteus

Pyrimidine dimer formation in response to UV radiation is governed by the thymine content of the potential dimer and the two flanking nucleotides. An enzymatic activity can be purified from Micrococcus luteus that cleaves the N-glycosyl bond between the 5' pyrimidine of a dimer and the corresponding sugar without rupture of a phosphodiester bond. We propose that strand scission at a dimer site by the M. luteus enzyme requires two activities, a pyrimidine dimer DNA-glycosylase and an apyrimidinic/apurinic endonuclease.     

Protection against malaria by immunization with plasmid DNA encoding cholera toxin B subunit andPlasmodium falciparum epitopes

Cholera toxin B subunit is a good carrier protein and an effective adjuvant which can boost both cellular and humoral immunity. DNA fragments encoding B cell, Th cell and CTL epitopes of P. falciparum CS, MSA-1, MSA-2 and RESA antigens were cloned down-stream of cholera toxin B subunit gene in the same reading frame. Another modification using IL2 as adjuvant was also made. High titer of anti-malaria epitopes antibodies and strong cellular immunogenicity were elicited after Balb/c mice were immunized three times with 100 μg recombinant plasmid DNA dissolved in 100 μL PBS. 200 vaccinees were challenged with mouse Plasmodium yoelli to investigate if cross protection existed. The protective efficacy was about 50%. And it is found that the protective efficacy is correlated with CTL activity which was considered to be the primary effects of anti-sporozoite protective immunity. Better results might be expected when the DNA vaccine candidates were applied to primates.     

DNA bending induced by cruciform formation

Cruciform structures in DNA are of considerable interest, both as extreme examples of sequence-dependent structural heterogeneity and as models for four-way junctions such as the Holliday junction of homologous genetic recombination. Cruciforms are of lower thermodynamic stability than regular duplex DNA, and have been observed only in negatively supercoiled molecules, where the unfavourable free energy of formation is offset by the topological relaxation of the torsionally stressed molecule. From an experimental viewpoint this can be a disadvantage, as cruciform structures can be studied only in relatively large supercoiled DNA circles, and are destabilized when a break is introduced at any point. We therefore set out to construct a pseudo-cruciform junction--by generating hereroduplex formation between two inverted repeat sequences. Stereochemically, this should closely resemble a true cruciform but remain stable in a linear DNA fragment. We have now created such a junction and find that it has the expected sensitivities to endonucleases. These DNA fragments exhibit extremely anomalous gel electrophoretic mobility, the extent of which depends on the relative position of the pseudo-cruciform along the length of the molecule. Our results are very similar to those obtained by Wu and Crothers using kinetoplast DNA, and we conclude that the pseudo-cruciform junction introduces a bend in the linear DNA molecule.     

微量Se保护细胞膜和细胞内DNA免受~1O_2损伤的研究

本文利用亚甲蓝光敏化产生~1O_2,对~1O_2的荧光检测法做了研究;利用滤膜过滤法研究了硒化合物保护NIH小鼠肾细胞DNA免受~1O_2损伤的作用;利用~3H-TdR掺入法研究了Na_2SeO_3对~1O_2抑制Wistar大鼠骨髓细胞DNA合成的作用;利用TBA法检测细胞脂质过氧化作用,观察到Na_2SeO_3抑制~1O_2诱导的细胞脂质过氧化的作用.Na_2SeO_3起保护作用的最适宜浓度范围为0.01～1.00μmol/L,高浓度的Na_2SeO_3表现出毒性作用.     

求解接点网络问题的DNA算法

利用DNA的二级结构——发卡构形，给出了求解接点网络问题的DNA算法．首先用DNA分子编码接点网络问题，然后利用DNA分子的自组装和形成二级结构的能力来求解问题．算法具有自动化实现计算的特点，计算所需的实验操作比Lipton提出的算法少，同时计算所需的DNA量也比Lipton提出的算法少．     

CpG DNA enhances the immune responses elicited by the DNA vaccine against foot-and-mouth disease virus in guinea pigs

CpG DNA is DNA sequence that has immune stimulatory effects. Several lines of investigation over the past few years indicate that CpG DNA plays an important role in the induction of immune responses to DNA vaccines. In this study, CpG DNA-containing synthetic oligodeoxynucleotide (CpG-ODN) was cloned into the eukaryotic expression plasmid encoding a fusion protein containing b- galactosidase from E. coli and immunogenic epitopes of foot- and-mouth disease virus (FMDV) type O, and the immune responses induced by the plasmid were assayed. The results showed that guinea pigs immunized with the recombinant plasmid containing CpG-ODN generated a higher level of FMDV-neutralizing antibody and a stronger T cell proliferative response and protection against viral challenge than those receiving the plasmid containing no CpG-ODN. Our study demonstrated that it is an effective route to enhance the efficacy of DNA vaccines by inserting exogenous CpG DNA into the plasmids, and the DNA vaccine developed here is a promising candidate to prevent FMDV infection.     

Telomeric DNA dimerizes by formation of guanine tetrads between hairpin loops

The telomeric ends of eukaryotic chromosomes are composed of simple repeating sequences in which one DNA strand contains short tracts of guanine residues alternating with short tracts of A/T-rich sequences. The guanine-rich strand is always oriented in a 5'-3' direction towards the end of the chromosome and is extended to produce a 3' overhang of about two repeating units in species where the telomeric terminus is known. This overhang has been implicated in the formation of several unusual intra-and intermolecular DNA structures, although none of these structures has been characterized fully. We now report that oligonucleotides encoding Tetrahymena telomeres dimerize to form stable complexes in solution. This salt-dependent dimerization is mediated entirely by the 3'-terminal telomeric overhang (TT-GGGGTTGGGG) and produces complexes in which the N7 position of every guanine in the overhangs is chemically inaccessible. We therefore propose that telomeric DNA dimerizes by hydrogen bonding between two intramolecular hairpin loops, to form antiparallel quadruplexes containing cyclic guanine base tetrads. These novel hairpin dimers may be important in telomere association and recombination and could also provide a general mechanism for pairing two double helices in other recombinational processes.     

Protection of cottontop tamarins against Epstein-Barr virus-induced malignant lymphoma by a prototype subunit vaccine

Epstein-Barr (EB) virus is one of the five herpesviruses of man. Strong links between this agent and the chain of events causing two human cancers, endemic Burkitt's lymphoma and undifferentiated nasopharyngeal carcinoma, have long been evident (reviewed in ref. 1). Because of this, and because of the very high incidence of nasopharyngeal carcinoma in certain large populations, it was suggested in 1976 that a vaccine should be developed against EB virus to prevent infection and thereby reduce tumour incidence amongst those at risk. The virus-determined membrane antigen (MA) was proposed as immunogen because it was known to elicit naturally occurring virus-neutralizing antibodies in man and because analogous antigens had been shown to act as effective experimental vaccines for preventing the herpesvirus-induced lymphomas of Marek's disease in chickens. Progress has been achieved in defining, quantifying and preparing MA molecules, and in enhancing their immunogenicity; a sensitive assay for antibodies to MA has been elaborated. Here we report that isolated cell membranes expressing MA, or purified MA glycoprotein of relative molecular mass (Mr) 340,000 (gp340), have been used to vaccinate cottontop tamarins (Saguinus oedipus oedipus), and that animals receiving either preparation were protected against the effects of a 100% tumour-inducing challenge dose of EB virus.     

Chiral recognition in dimerization of adsorbed cysteine observed by scanning tunnelling microscopy

Stereochemistry plays a central role in controlling molecular recognition and interaction: the chemical and biological properties of molecules depend not only on the nature of their constituent atoms but also on how these atoms are positioned in space. Chiral specificity is consequently fundamental in chemical biology and pharmacology and has accordingly been widely studied. Advances in scanning probe microscopies now make it possible to probe chiral phenomena at surfaces at the molecular level. These methods have been used to determine the chirality of adsorbed molecules, and to provide direct evidence for chiral discrimination in molecular interactions and the spontaneous resolution of adsorbates into extended enantiomerically pure overlayers. Here we report scanning tunnelling microscopy studies of cysteine adsorbed to a (110) gold surface, which show that molecular pairs formed from a racemic mixture of this naturally occurring amino acid are exclusively homochiral, and that their binding to the gold surface is associated with local surface restructuring. Density-functional theory calculations indicate that the chiral specificity of the dimer formation process is driven by the optimization of three bonds on each cysteine molecule. These findings thus provide a clear molecular-level illustration of the well known three-point contact model for chiral recognition in a simple bimolecular system.     

Single-site enzymatic cleavage of yeast genomic DNA mediated by triple helix formation.

Physical mapping of chromosomes would be facilitated by methods of breaking large DNA into manageable fragments, or cutting uniquely at genetic markers of interest. Key issues in the design of sequence-specific DNA cleaving reagents are the specificity of binding, the generalizability of the recognition motif, and the cleavage yield. Oligonucleotide-directed triple helix formation is a generalizable motif for specific binding to sequences longer than 12 base pairs within DNA of high complexity. Studies with plasmid DNA show that triple helix formation can limit the operational specificity of restriction enzymes to endonuclease recognition sequences that overlap oligonucleotide-binding sites. Triple helix formation, followed by methylase protection, triple helix-disruption, and restriction endonuclease digestion produces near quantitative cleavage at the single overlapping triple helix-endonuclease site. As a demonstration that this technique may be applicable to the orchestrated cleavage of large genomic DNA, we report the near quantitative single-site enzymatic cleavage of the Saccharomyces cerevisiae genome mediated by triple helix formation. The 340-kilobase yeast chromosome III was cut uniquely at an overlapping homopurine-EcoRI target site 27 base pairs long to produce two expected cleavage products of 110 and 230 kilobases. No cleavage of any other chromosome was detected. The potential generalizability of this technique, which is capable of near quantitative cleavage at a single site in at least 14 megabase pairs of DNA, could enable selected regions of chromosomal DNA to be isolated without extensive screening of genomic libraries.