Hybridization of two oligodeoxynucleotides to both strands of an RNA hairpin structure increases the efficiency of RNA‐DNA duplex formation

Malygin, Alexey A.; Карпова, Г. Г.; Westermann, Peter

doi:10.1016/0014-5793(96)00798-3

Cited by 9 publications

(7 citation statements)

References 19 publications

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“…Partial alkaline hydrolysis (Fig. 7A, lane 1) was identical to the control (lane 2) up to the phosphodiester bond secondary structure (Malygin et al, 1996). Therefore, alkylation of 4.5S RNA by ON1-3′-R was studied using a large excess of 3′ to G58.…”

Section: Synthesis Of P-(mentioning

confidence: 85%

“…Hybridization of lent RNA-DNA complexes were hydrolyzed by incubation in 50 mM NaHCO 3 , pH 8.9, in the presence of 200 µg/ml E. coli 4.5S RNA with ON2 is restricted by the RNA secondary structure and the extent of hybridization has been determined before total tRNA at 90°C for 15 min. Hydrolysis products were separated on a 10% polyacrylamide gel and compared with a (Malygin et al, 1996). To optimize hybridization and maximize the alkylation reaction, ratios between 5′-R-ON2 and 4.5S RNA 'ladder' obtained by degradation of unmodified 4.5S RNA.…”

Section: Synthesis Of P-(mentioning

confidence: 99%

“…The most informative approaches are reactions of RNA molecules carrying indivi-formation of a ternary complex up to 10-fold (Malygin et al, 1996). Here, we use chemically reactive oligodeoxynucleotide dual activated bases.…”

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Site‐specific modification of 4.5S RNA apical domain by complementary oligodeoxynucleotides carrying an alkylating group

Bulygin

Malygin

Карпова

et al. 1998

European Journal of Biochemistry

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Site-specific alkylation of RNA by reactive oligodeoxynucleotides provides structural information and represents the first step towards the design of RNA derivatives to be used for functional studies. Specific alkylation of 4.5S RNA at G53, the first base of the apical tetraloop, was achieved by incubation with oligodeoxynucleotide ON2, complementary to nucleotides 38Ϫ53, which carries a p- benzylamidophosphate group at the 5′ end. Alkylation efficiency was increased by a factor of 6, without alteration of specificity, in the presence of a helper oligodeoxynucleotide, ON1, complementary to nucleotides 58Ϫ71 of the opposite strand of the RNA helix. A second reactive oligodeoxynucleotide, ON1-3′-R, was obtained by attaching the alkylating group to the 3′ end of ON1. ON1-3′-R was able to modify G58. In the presence of ON2 as a helper oligodeoxynucleotide, the specificity of ON1-3′-R changes and efficient alkylation of nucleotides G54, A56 and G57 of the apical region of 4.5S RNA was observed.Keywords : Escherichia coli 4.5S RNA; RNA secondary structure ; site-specific modification ; reactive oligodeoxynucleotide.Interactions of RNA with other macromolecules are studied, complementary to the apical stem-loop structure of 4.5S RNA of Escherichia coli were shown to stimulate synergistically the among others, by cross-linking approaches. The most informative approaches are reactions of RNA molecules carrying indivi-formation of a ternary complex up to 10-fold (Malygin et al., 1996). Here, we use chemically reactive oligodeoxynucleotide dual activated bases. Derivatives of small RNAs can be synthesized chemically whereas methods to modify large RNA mole-derivatives bearing a p-(N-2-chloroethyl-N-methylamino)benzylamidophosphate group at either the 5′ or 3′ ends. After hybridcules at specific sites are still a challenging task. A very promising approach involves chemically reactive derivatives of anti-ization with 4.5S RNA, site-specific alkylation is observed.Upon combination with a second unmodified oligodeoxynucleosense oligonucleotides, among which phosphamide derivatives with a terminal alkylation group have proven to be most useful tide complementary to the opposite RNA strand, alkylation efficiencies are improved. In addition, a different hybrid structure (Zenkova et al., 1990(Zenkova et al., , 1995Venkstern et al., 1990). They are able to modify any nucleotide residue in RNA (except uridine) was observed in the presence of an oligodeoxynucleotide complementary to bases 38Ϫ53 of 4.5S RNA (ON2) as a helper neighboring the oligonucleotide 5′ or 3′ ends (Knorre et al., 1986). By this method, aminobenzyl groups are introduced, oligodeoxynucleotide which resulted in alkylation of another set of nucleotides within the apical domain of 4.5S RNA. which in turn can be converted into site-specific, photoreactive derivatives of RNAs. Modification efficiencies largely depend on the extent of hybridization between the target nucleic acid and the complementary oligodeoxynucleotide that often is inhib-MATERIALS AND METHODS ited...

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Section: Synthesis Of P-(mentioning

confidence: 85%

Section: Synthesis Of P-(mentioning

confidence: 99%

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Site‐specific modification of 4.5S RNA apical domain by complementary oligodeoxynucleotides carrying an alkylating group

Bulygin

Malygin

Карпова

et al. 1998

European Journal of Biochemistry

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“…To increase the yield of RNA modification at the desired site in the cases when target sequences are significantly involved in the intra-RNA interactions, an approach was suggested that utilized helper oligodeoxyribonucleotides facilitating unfolding of RNA structure in the target sequence region. With E. coli 4.5S RNA as a model target RNA, it was demonstrated that two oligonucleotides complementary to the opposite strands of the RNA apical stem-loop structure stimulate synergistically the formation of a ternary complex up to 10-fold [44]. Application of this approach with -NH-CH 2 RCl derivatives of deoxy-oligomers and the same target RNA showed that helpers can increase efficiency of RNA alkylation by a factor of 6; it was also found that helper oligomer can alter specificity of target RNA alkylation.…”

Section: Approach For Site-specific Introduction Of Amino Linkers mentioning

confidence: 99%

General Approach for Introduction of Various Chemical Labels in Specific RNA Locations Based on Insertion of Amino Linkers

Graifer

Карпова

2013

Molecules

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Introduction of reporter groups at designed RNA sites is a widely accepted approach to gain information about the molecular environment of RNAs in their complexes with other biopolymers formed during various cellular processes. A general approach to obtain RNAs bearing diverse reporter groups at designed locations is based on site-specific insertion of groups containing primary aliphatic amine functions (amino linkers) with their subsequent selective derivatization by appropriate chemicals. This article is a brief review on methods for site-specific introduction of amino linkers in different RNAs. These methods comprise: (i) incorporation of a nucleoside carrying an amino-linker or a function that can be substituted with it into oligoribonucleotides in the course of their chemical synthesis; (ii) assembly of amino linker-containing RNAs from short synthetic fragments via their ligation; (iii) synthesis of amino linker-modified RNAs using T7 RNA polymerase; (iv) insertion of amino linkers into unmodified RNAs at functional groups of a certain type such as the 5'-phosphates and N7 of guanosine residues and (v) introduction of an amino linker into long highly structured RNAs exploiting an approach based on sequence-specific modification of nucleic acids. Particular reporter groups used for derivatization of amino linker-containing RNAs together with types of RNA derivatives obtained and fields of their application are presented.

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“…Instead of modifying the oligonucleotide funtionalities, we hypothesized that novel ligands could be introduced to enhance the stability of the RNA-DNA heteroduplexes. The study of heteroduplexes have been active over the last few years, [19][20][21][22][23][24] and the generation of tight-binding ligands against heteroduplex may offer a more efficient antisense system without the need to introduce modified oligonucleotide.…”

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A comparative binding study of BIV Tat peptide against its TAR RNA duplex, RNA–DNA heteroduplex and DNA duplex

Tok

2005

Bioorganic & Medicinal Chemistry Letters

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Hybridization of two oligodeoxynucleotides to both strands of an RNA hairpin structure increases the efficiency of RNA‐DNA duplex formation

Cited by 9 publications

References 19 publications

Site‐specific modification of 4.5S RNA apical domain by complementary oligodeoxynucleotides carrying an alkylating group

Site‐specific modification of 4.5S RNA apical domain by complementary oligodeoxynucleotides carrying an alkylating group

General Approach for Introduction of Various Chemical Labels in Specific RNA Locations Based on Insertion of Amino Linkers

A comparative binding study of BIV Tat peptide against its TAR RNA duplex, RNA–DNA heteroduplex and DNA duplex

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