Biomedical subjects

D Cech

Publications and source records attributed to D Cech.

27 records · Page 2Linked to original sources

PubMed1986

Chemical synthesis of octadeoxyribonucleotides containing sequence-specific 3'-deoxy-3'-chlorouridine residues and 2'-5' internucleotide linkages by the phosphite chloridite approach.

3'-Deoxy-3'-chlorouridine has been chemically incorporated into two octanucleotides at various positions using in situ prepared phosphite chloridite intermediates. The behaviour of these modified selfcomplementary DNA fragments containing 2'-5' internucleotide bonds in the neighbourhood of 3'-deoxy-3'-chlororibose in respect to chemical sequencing, polynucleotide kinase and T4 DNA ligase has been studied.

Base Sequence↗

PubMed1985

Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments. VI. The binding and cleavage of substrates containing nucleotide analogs.

The present study deals with the binding and cleavage by EcoRII endonuclease of concatemer DNA duplexes containing EcoRII recognition sites (formula; see text) in which dT is replaced by dU or 5-bromodeoxyuridine, or 5'-terminal dC in the dT-containing strand is methylated at position 5. The enzyme molecule is found to interact with the methyl group of the dT residue of the DNA recognition site and to be at least in proximity to the H5 atom of the 5'-terminal dC residue in dT-containing strand of this site. Modification of any of these positions exerts an equal effects on the cleavage of both DNA strands. Endonuclease EcoRII was found to bind the substrate specifically. At the same time modification of the bases in recognized sequence may result in the formation of unproductive, though stable, enzyme-substrate complexes.

Base Sequence↗

PubMed1985

[Chemical synthesis, isolation and sequencing of tetradecadeoxyribonucleotides containing the modified bases 5-fluorouracil and 5-methylcytosine].

It is reported on phosphate-triester synthesis, isolation and sequencing of five tetradecadeoxyribonucleotides containing the modified bases 5-fluorouracil and 5-methylcytosine. The modified nucleotides can be exactly determined in the oligonucleotide chain by solid-phase MAXAM/GILBERT-sequencing. By combination of the oligonucleotides new hemi-modified DNA-duplexes were constructed which serve as model compounds for investigating the interaction with the EcoRII restriction and modification enzymes.

5-Methylcytosine↗

PubMed1983

Mass spectrometric monitoring of solid phase phosphate triester synthesis of DNA fragments.

A new and convenient pyrolysis mass spectrometric approach for monitoring solid phase phosphate triester synthesis of oligodeoxyribonucleotides in 5'-3' direction has been developed. The base-typical fragment ions produced by pyrolysis at 280 degrees C of the polymer-bound oligonucleotide triesters in the mass spectrometer permit the analytical monitoring of DNA chain growth, using simple mathematical operations. The base composition can be determined directly from the polymer. In addition, minor nucleosides can easily be detected.

Base Sequence↗

PubMed1983

Antiherpes activity of (E)-5-(2-bromovinyl)- and 5-vinyl-1-beta-D-arabinofuranosyluracil and some other 5-substituted uracil arabinosyl nucleosides in two different cell lines.

Of a series of 5-substituted 1-beta-D-arabinofuranosyluracil (5-X-araU) analogues, (E)-5-(2-bromovinyl)-araU(BrVaraU) and 5-vinyl-araU (VaraU) were the most potent inhibitors of plaque formation by two herpes simplex virus type 1 (HSV-1) strains in human embryonic lung fibroblast (HELF) cell cultures. They were not only more active than 5-methyl-araU (MaraU, araT) and 5-ethyl-araU (EaraU), but even more than 1000 times more potent than the 5-fluoro, 5-iodo, 5-formyl and 5-trifluoromethyl (FaraU, IaraU, faraU, CF3araU) analogues. BrVaraU and VaraU were superior to 9-(2-hydroxyethoxymethyl)guanine (Acyclovir, ACV) and comparable in potency with 2'-fluoro-5-iodoaracytosine (FIAC) and 2'-fluoro-5-methylarauracil (FMAU). Their anti-HSV-1 potency was surpassed only by (E)-5-(2-bromovinyl)-2'-deoxyuridine (BrVUdR). Surprisingly, in a HSV-1 plaque inhibition assay in African green monkey kidney (Vero) cells, BrVaraU and VaraU were nearly 100 times active or even inactive. In contrast, the antiherpes activity of ACV, FIAC, FMAU and BrVUdR differed only marginally in the two cell lines. The following order of (decreasing) activity against HSV-2 in HELF cells was found: FIAC = FMAU greater than MaraU (araT) greater than ACV greater than VaraU greater than BrVUdR greater than CF3araU greater than IaraU greater than FaraU = Eara U greater than BrVaraU greater than araU greater than faraU. When deoxyribose is replaced by arabinose in 5-X-UdR analogues, a slight increase in anti-HSV-1-77 activity was observed for the 5-vinyl or 5-ethyl substituent, whereas the other 5-X-araU nucleosides were two to more than 100 times less active than their deoxyribosyl counterparts. However, the sugar exchange led to a strong reduction in anti-HSV-2 activity regardless of the 5-substituent.

Animals↗

PubMed1977

Fluorination of 6-methyluracil and its nlcleosides.

6-Methyluracil and its perbenzoylated 1-(beta-D-ribofuranosyl) and 1-(2-deoxy-beta-D-ribofuranosyl) derivatives afford on treatment with elemental fluorine in acetic acid solutions, the corresponding derivatives of 5-fluoro-6-methyluracil and 5-fluoro-6-fluoromethyluracil. The free nucleosides have been obtained from the protected derivatives by methanolysis. The CH2F linkage in 5-fluoro-6-fluoromethyluracil derivatives is stable towards hydrolysis and nucleophilic agents.

Chemical Phenomena↗

PubMed1975

Synthesis of substituted 5-fluoro-5,6-dihydropyrimidines.

The reaction of 5-substituted uracils with fluorine in acetic acid and other solvents and the following treatment with different alcohols yielded the corresponding 5-fluoro-5,6-substituted-5,6-dihydropyrimidines. Thymine gave 5-fluoro-5-methyl-6-alkoxy-5,6-dihydropyrimidines. 5-Halogeno uracils and 5-nitrol uracil were converted into 5-fluoro-5-halogeno-6-hydroxy-5,6-dihydropyramidines and the 5-nitroanalogue, respectively. The structures of the compounds were confirmed by mass spectrometry.

Fluorine↗

PubMed1975

A facile synthesis of 5-(perfluoroalkyl)-pyrimidines.

In the paper a synthetic two stage procedure is described for the preparation of perfluoroalkylated derivatives of uracil and its nucleosides. Using copper bronze a perfluoroalkyl-copper-complex is formed from perfluoralkyl iodides in polar aprotic solvents, such as DMSO, and under inert conditions. The reaction of this complex with uracil, uridine and 2-deoxyuridine leads to the corresponding 5-substituted perfluoralkyl derivatives. It is shown by mass spectra that the substitution always takes place at the 5-position of the pyrimidine. The chemical and physical properties of the formed compounds are described.

Deoxyuridine↗

PubMedDate not supplied

Versatile procedure of multiple introduction of 8-aminomethylene blue into oligonucleotides.

The coupling of 8-aminomethylene blue to oligonucleotides via poly-L-glutamic acid linker using carboxy-anchor groups will be described. The introduction of carboxy-anchor groups into oligonucleotides proceeds both during automated synthesis using 6-(ethoxycarbonyl)hexyl 1-O-phosphoramidite and by reaction of 5'-amino-functionalized oligonucleotides with succinic anhydride. O-(N-Succinimidyl)-1,1,3,3-tetramethyluronium tetrafluoroborate was used as activating reagent for binding of poly-L-glutamic acid to the carboxylated oligonucleotides. The successful 5'-carboxylation and poly-L-glutamic acid coupling were proven both by polyacrylamide gel electrophoreses and HPLC. 8-Aminomethylene blue in its leucoform was covalently coupled to the oligonucleotides in the presence of water soluble carbodiimide.

Base Sequence↗