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M Zenkova

Publications and source records attributed to M Zenkova.

13 recordsLinked to original sources
PubMed2006

Artificial ribonucleases: from combinatorial libraries to efficient catalysts of RNA cleavage.

Combinatorial libraries of small organic compounds capable of cleaving RNA were synthesized. The compounds contain benzene ring substituted with two residues of bis quaternary salt of diazabicyclo[2.2.2]octane (DABCO) bearing hydrophobic fragments of different length and structure, attached to DABCO at the bridge position. These compounds, lacking traditional functionalities involved in transesterification reaction, exhibit pronounced RNA cleavage activity. To identify the most active artificial ribonucleases, sublibraries and truncated libraries, containing compounds lacking one of substituents were synthesized. Analysis of ribonuclease activity of truncated libraries resulted in identification of the most active compounds, which are characterized by the presence of at least one long oligomethylene substituent.

Alkanes↗
PubMed2006

Binary hammerhead ribozymes with improved catalytic activity.

A new design of binary hammerhead ribozymes displaying high catalytic activity and nucleolytic stability is described. These catalytic structures consist of two partially complementary oligoribonucleotides, capable of assembling into the hammerhead-like structure without tetraloop II on binding to the RNA target. A series of these binary ribozymes targeting the translation initiation region of multiple drug resistance gene mdr1 mRNA was synthesized and assessed in terms of catalytic activity under single and multiple reaction turnover conditions. Enhanced nuclease resistance of the binary ribozymes was achieved by incorporation of 2'-modified nucleotides at selected positions, along with addition of a 3'-3'-linked thymidine cap. The new binary ribozymes exhibit higher RNA cleavage activity than their full-length analogs because of faster dissociation of cleavage products. Furthermore, an excess of one of the ribozyme strands provides the possibility to unfold structured regions of the target RNA and facilitate productive complex formation.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2005

Modified binary hammerhead ribozymes with high catalytic activity.

A series of binary hammerhead ribozymes was designed and assessed in terms of cleavage activity and nuclease resistance. Enhanced nuclease resistance of binary ribozymes was achieved by incorporation of Z-modified nucleotides at the selective positions along with addition of 3'-3-linked thymidine cap. These modified binary ribozymes efficiently cleave 190-nucleotides long MDR1 mRNA fragment and display catalytic activity much higher then respective full-length analogs.

Base Composition↗
PubMed2004

The role of hydrophobic interactions in catalysis of RNA cleavage by 1,4-diazabicyclo[2.2.2]-octane based artificial ribonucleases.

Molecular interactions of RNA cleaving compounds-conjugates of 1,4-diazabicyclo[2.2.2.]octane substituted at the bridge position with tetradecamethylene fragment and imidazole were investigated using light scattering and small angle x-ray scattering methods. The compounds are known to efficiently cleave RNA and one source of the activity could result from micellar catalysis. It was found that the compounds indeed are capable of forming complex aggregates in solution. However, maximal efficacy of RNA cleavage by the conjugates is observed at concentrations well below the concentration required for micelle formation.

Catalysis↗
PubMed2004

Ribonuclease activity of cationic structures conjugated to lipophilic groups.

Cationic compounds containing benzene ring substituted with the bis-quaternary salt of diazabicyclo[2.2.2]octane (DABCO) bearing a polymethylene fragment at the bridge positions display ribonuclease activity. Efficacy of the catalysis is affected by geometry of the cationic structures and the size of the attached aliphatic fragment. The cleavage occurs primarily within CA sequences. The compounds do not possess tradition groups participating in the transesterification step of RNA cleavage reaction, therefore a speculative mechanism of cleavage could be inducing a conformational stress on the RNA sugar phosphate backbone providing fragility to phosphodiester bonds.

Autoradiography↗
PubMed2001

5'-bis-pyrenylated oligonucleotides displaying excimer fluorescence provide sensitive probes of RNA sequence and structure.

Oligonucleotide conjugates bearing two pyrene residues attached to 5'-phosphate through a phosphoramide bond were synthesised. Fluorescence spectra of the conjugates show a peak typical of monomer emission (lambda(max) 382 nm) and a broad emission peak with lambda(max )476 nm, which indicates the excimer formation between the two pyrene residues. Conjugation of these two pyrene residues to the 5'-phosphate of oligonucleotides does not affect the stabilities of heteroduplexes formed by conjugates with the corresponding linear strands. A monomer fluorescence of the conjugates is considerably affected by the heteroduplex formation allowing the conjugates to be used as fluorescent hybridisation probes. The 5'-bis-pyrenylated oligonucleotides have been successfully used for investigation of affinity and kinetics of antisense oligonucleotides binding to the multidrug resistance gene 1 (PGY1/MDR1) mRNA. The changes of excimer fluorescence of the conjugates occurring during hybridisation depended on the structure of the binding sites: hybridisation to heavily structured parts of RNA resulted in quenching of the excimer fluorescence, while binding to RNA regions with a loose secondary structure was accompanied by an enhancement of the excimer fluorescence. Potentially, these conjugates may be considered as fluorescent probes for RNA structure investigation.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Invasion of strongly binding oligonucleotides into tRNA structure.

Interaction of yeast tRNA(Phe) with oligodeoxyribonucleotides containing 5-methylcytosine, 2-aminoadenine, and 5-propynyl-2'-deoxyuridine was investigated. The modified oligonucleotides show increased binding capacity although the association rates are similar for the modified and natural oligonucleotides. The most pronounced increase in association constant (70 times) due to the incorporation of the strongly binding units was achieved in the case of oligonucleotide complementary to the sequence 65-76 of the tRNA(Phe).

2-Aminopurine↗
PubMed1995

A novel approach to introduce site-directed specific cross-links within RNA-protein complexes. Application to the Escherichia coli threonyl-tRNA synthetase/translational operator complex.

We describe a methodology which allows the introduction of a photoactivatable azido group at specific internal positions of any RNA in order to identify the neighboring elements of an interacting protein. The first step involves site-directed modification of the target RNA with an antisense oligodeoxyribonucleotide bearing, at its 3' or 5' phosphate, a 4-[-N-(2-chloroethyl)-N-methylamino]benzylmethylamino group. Position N7 of a guanine residue located in the close vicinity of the hybrid is the main target for alkylation. The antisense oligodeoxyribonucleotide is then removed by acidic pH treatment and a photoreactive reagent (2,4-dinitro-5-fluorophenylazide) is condensed to the modified nucleotide. This method was used to induce specific cross-links between Escherichia coli threonyl-tRNA synthetase and the leader region of threonyl-tRNA synthetase mRNA, which is involved in translational feedback regulation. Control experiments revealed that the modification affects neither the structure of the mRNA nor the interaction with the enzyme. More than 50% of the modified mRNA complexed with threonyl-tRNA synthetase can be cross-linked to the enzyme, depending on the nucleotide modified.

Alkylation↗
PubMedDate not supplied

Interaction of complementary oligonucleotides with the 3'-end of yeast tRNA(Phe).

Interaction of yeast tRNA(Phe) with oligodeoxyribonucleotides (ONs), complementary to the nucleotides 62-76 was investigated. Results of gel-mobility shift assay and RNase A probing evidence that the ONs containing the sequence complementary to the tRNA ACCA end can easily invade the hairpin structure under physiological conditions. The limiting step of association process is the tRNA unfolding.

Nucleic Acid Conformation↗
PubMedDate not supplied

Mechanism and specificity of RNA cleavage by chemical ribonucleases.

Cleaving of model RNA substrates by chemical ribonucleases constructed by conjugation of 1,4 diazabicyclo[2,2,2]octane with histamine and histidine was investigated. Similarly to RNase A, the chemical RNases produce fragments with 5' hydroxy-group and 3'-cyclophosphate. The cleavage occurs as the catalytic reaction: more than 150 phosphodiester bonds in RNA can be cleaved by one molecule of RNase mimic.

Base Sequence↗
PubMedDate not supplied

2'-modified oligonucleotides from methoxyoxalamido and succinimido precursors: synthesis, properties, and applications.

Synthesis of 2'-modified oligonucleotides from 2'-methoxyoxalamido (MOX) and 2'-succinimido (SUC) precursors is described. Their physical and biochemical properties were assessed. Synthesized oligonucleotides were used as primers in advanced DNA sequencing protocols. An example of sequencing directly off genomic DNA template without prior cloning or PCR amplification is presented.

Base Sequence↗
PubMedDate not supplied

5'-bis-pyrenylated oligonucleotides display enhanced excimer fluorescence upon hybridization with DNA and RNA.

A simple one-step procedure was applied for synthesis of oligonucleotide conjugates bearing two pyrene residues at the 5'-phosphate of oligonucleotide. Excimer fluorescence intensity of the conjugates is highly sensitive to duplex formation: binding of the bis-pyrenylated oligonucleotides to their DNA and RNA targets leads 10-fold increase of fluorescence. The data show that excimer fluorescence intensity of the conjugates depends linearly on the concentration of target DNA and permits quantification of DNA in solution.

Base Sequence↗