Nucleic acids symposium series (2004)最新文献

DNA damage was monitored by fluorescent microscopy observations of DNA fluorescent images after hydrodynamic stretching on a microscope glass. DNA double-strand breaks lead to a decrease of the average length of observed fluorescent DNA molecules. Compared to conventional methods such as electrophoresis, the proposed method allows for the analysis of the DNA damage at very low DNA breaking frequency. In particular, this method was used to study DNA damage by weak UV irradiation in solutions of quantum dots.

Introduction of therapeutic genes into the cells of an organism in a safe and efficient way has become a challenging task in non-viral mediated gene therapy. Here, branched polyethylenimine (bPEI, 25 kDa) was converted into nanoparticles through electrostatic interactions with anionic polysaccharides (e.g. alginic acid, Al and hyaluronic acid, HA). A small library of PEI-Al and PEI-HA nanoparticles was synthesized by varying the amounts of anionic polysaccharides and evaluated in terms of their size, surface charge, cytotoxicity, transfection efficiency, etc. Both the series of nanoparticles exhibited higher cell viability and transfection efficiency as compared to native PEI and the standard transfection reagents. In vivo targeting efficacy of PEI-HA(4.6%) nanoparticles was examined in tumor induced mice.

HAC1 is a transcription factor related to Unfolded Protein Response (UPR) signaling in yeast. Processing of HAC1 mRNA on Endoplasmic reticulum (ER) plays a key role in UPR signaling pathway, but the recognition mechanism of HAC1 mRNA by processing enzyme Ire1p is still unclear. Here, the solution structure of HAC1 mRNA was investigated by Nuclear Magnetic Resonance (NMR) spectroscopy, focusing on the structure of the recognition site of Ire1p in HAC1 mRNA. From the NOESY spectrum, imino proton signals of 5' processing regions of HAC1 mRNA were assigned and it was found that this region forms the stem-loop structure.

Polyamines, especially branched polyamines such as tetrakis(3-aminopropyl)ammonium (Taa), stabilize the tertiary structure of RNA molecules. In this study, we examined the polyamine binding site of the HIV-1 dimerization initiation site (DIS) in the kissing-loop dimer by the docking simulation. It was found that Taa binds predominantly to the kissing loop interaction site of DIS.

The intermolecular DNA quadruplex is attractive materials for design of nanomolecular constructs and machines. This folding kinetics is, however, slow, hampering its application to these materials. We have reported that the cationic comb-type copolymer accelerated duplex and triplex formation. In this study, the effect of the copolymer on the intermolecular quadruplex folding was investigated. The copolymer was found to accelerate the association of intermolecular quadruplex by three-orders. Furthermore, the copolymer induced the strand exchange reaction between intermolecular quadruplex and its single-stranded counterparts. We suggested that the copolymer acts as a nucleic acid chaperone for the intermolecular quadruplex.

DNA duplexes containing diaminostilbene (DAS) as a photoinduced electron donor were synthesized to investigate mechanisms of electron injection into DNA and the succeeding electron transfer in the duplexes. DAS-Capped hairpin DNA showed a high structural stability thereby attains large interaction between DAS and the terminal base pair. DAS-Tethered DNA by a single linker at the end of the duplex was also synthesized and the yields of photoinduced electron transfer through mismatched base pairs were quantified. Both duplexes showed similar electron transfer efficiencies depending on the base pairs, which suggests DAS stacks well on the "pi-way" of the duplex DNA.

We present a novel photoelectrochemical approach for discriminating between cytosine and 5-methylcytosine in DNA when used in combination with enzymatic digestion. A photosensitizer-linked DNA duplex bearing 5-methylcytosine or cytosine at a given restriction site of the strand was immobilized on a gold electrode and digested with enzyme, and the photocurrent response was measured. We observed high photocurrent density that was selective for the methylated duplex.

Next generation 4'-selenonucleosides as potential antiviral and antitumor agents were synthesized using a Pummerer type condensation as a key step.

Twisted intercalating nucleic acids form stable triplexes with polypurine tracts of double-stranded DNA. Their affinity depends on their length, primary structure and base contents, parallel or antiparallel orientation of oligonucleotides respectively to DNA, number of TINA residues and their relative positions. Basing on parallel CT, GT and antiparallel GT triplex-forming 16-mer oligonucleotides targeted to polypurine tract of HIV proviral DNA, we synthesized eleven different oligonucleotides with 2-4 TINA insertions in different positions. Studies of their interaction with target duplex by gel shift, fluorescence spectroscopy, circular dichroism and thermal denaturation demonstrated that antiparallel GT oligonucleotides form more stable triplexes than parallel TC or TG ones. Two best candidates were selected for the further studies. The first one (5'-AGGGxGGGTTTxTGTTTT-3', Kd = 219 nM) contains only two TINA insertions and does not aggregate in non-denaturing conditions, in contrast to majority of other oligonucleotides.

A novel fluorescent oligonucleotide probe containing pyrene-labeled C8 alkylamino-substituted 2'-deoxyguanosine and a practically useful 3'- and 5'- ends-free self-quenched molecular beacons (MB) were designed. A unique MB detectable by pyrene excimer fluorescence was also demonstrated.