Nucleic acids research. Supplement (2001)最新文献

Application of 2-acetoxyethyl acetoxymethyl ether as a model of peracylated sugars (e.g. 1,2,3,5-tetra-O-acetyl-beta-D-ribofuranose) is a convenient way to study the mechanism and sequence of glycosyl exchange reactions in the nucleoside chemistry.

2',6'-Dideoxy-beta-D-hexopyranosyl pyrimidine nucleoside, which is a component of anticoccidal antibiotics--cytosaminomycins, was synthesized in a stereoselective manner via the intramolecular pyrimidine delivery method. Further glycosylaion of the 4'-position with a glycosyl fluoride gave the disaccharide nucleoside skeleton of the cytosaminomycins.

Heterocyclic amines (HCAs) are mutagenic compounds present in cooked meat and fish, and some of them are known to be carcinogenic. DNA adduct formation is now believed to be the initial critical step for carcinogenesis. HCAs are metabolically activated to form predominantly 2'-deoxyguanosine-C8 adducts (dG-C8 adduct) where the exocyclic N atom of activated compounds are covalently bound to the C8 position of dG. In order to prepare a high yield of dG-C8 adducts with HCAs, we tried to adapt the Buchwald-Hartwig arylamination reaction using 8-bromo-dG derivatives and HCAs. With the combined use of xantphos and Cs2CO3, we successfully obtained coupling compound derived from a carcinogenic HCA, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) at a 97% yield. Subsequent deprotection may lead to authentic dG-C8 adducts of several HCAs.

Perturbation of active site functional group pKas is an important strategy employed by protein enzymes to achieve catalysis. There is increasing evidence to indicate that RNAs also utilize functional group pKa perturbation for folding and reactivity. The two best candidates for a functionally relevant pKa perturbation are the N3 of C (pKa 4.2) and the N1 of A (pKa 3.5), either of which could be sufficiently raised to allow protonation near physiological pH. Here we report the synthesis and use of a series of alpha-phosphorothioate tagged cytidine and adenosine analogs whose altered pKas make it possible to efficiently detect functionally relevant protonation events by Nucleotide Analog Interference Mapping. This approach has been used to detect ionization events in several catalytic RNAs, including the group I intron, the Hepatitis Delta Virus (HDV), the hairpin and the Varkud Satellite (VS) ribozymes. The active site residue of the VS ribozyme appears to be ionized in the course of the reaction pathway, which may be indicative of a general acid or base mechanism for catalysis by this RNA.

The minisatellite DNA Pc-1 consists of tandem repeats of d(GGCAG). We previously reported that a d(GGCAG)n strand folds into an intramolecular quadruplex under physiological conditions and that during replication the progression of DNA polymerase is blocked by the quadruplex in vitro. Therefore, the formation of the quadruplex was supposed to be responsible for the hypermutable features of Pc-1. Then, we have identified proteins that bind to Pc-1, one of which is hnRNP A1. Here, we have demonstrated that hnRNP A1 destroys the quadruplex of Pc-1 on binding and abrogates the arrest of DNA polymerase at the repeat. Thus, hnRNP A1 functions as if it is a chaperon to assist Pc-1 DNA to form the proper folding suitable for replication. We have also found that hnRNP A1 and a related protein, hnRNP D, destroy the quadruplex of telomere DNA, which suggests the involvement of these proteins in telomere maintenance as DNA chaperons.

By using appropriately modified oligonucleotides, a gap-structure is formed in substrate DNA, and 2-6 ethylenediamine-N,N,N'-triacetate residues are placed near this gap. When these composites are treated with homogenous Ce(IV)/EDTA complex, the phosphodiester linkages in the gap-site are selectively hydrolyzed at much greater rates than achieved previously by using unmodified oligonucelotides.

2,6-Diamino-1,8-naphthyridine derivative (daNpt), which possesses hydrogen bonding groups in an alignment of donor-acceptor-acceptor-donor binds to a single nucleotide bulge in the duplex. The melting temperatures (Tm) of all duplexes containing a bulge were increased, especially for the cytosine (C) and thymine (T) bulges, in the presence of daNpt, whereas only a small increase of Tm was observed for the fully matched duplexes. It was suggested by pH dependency of Tm and UV spectra, that a protonation of daNpt should play an important role for the recognition of C and T bulges.

5,10,15,20-Tetra(N-methyl-pyridinium-4-yl)-21H,23H-porphyrin has shown to bind to the major groove of AT-rich DNA predominantly through electrostatic interaction between positively charged N-methyl pyridinium moieties of the porphyrin and negatively charged phosphodiester backbone. Solution structure of the complex between the porphyrin and a double-stranded DNA fragment has been inferred from the measurements of the mixing time-dependent intermolecular nuclear Overhauser effects (NOEs).

Naphthyl Red moiety, conjugated to DNA, shows distinct chromism by hybridization with its complementary DNA. Single-stranded DNA involving Naphthyl Red moiety exhibits orange color and has lambda max at 466 nm at pH 7.0. Absorption maximum shifts towards 545 nm by the presence of its complementary DNA, and accordingly color of the solution changes from orange to magenta.