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Remarkable acceleration of a DNA/RNA inter-strand functionality transfer reaction to modify a cytosine residue: the proximity effect via complexation with a metal cation.

Jitsuzaki D, Onizuka K, Nishimoto A, Oshiro I, Taniguchi Y, Sasaki S - Nucleic Acids Res. (2014)

Bottom Line: We have recently developed a new strategy for the in situ modification of RNA based on the functionality transfer reaction between an oligodeoxynucleotide probe and an RNA substrate. 2'-Deoxy-6-thioguanosine (6-thio-dG) was used as the platform to anchor the transfer group.It was demonstrated that the (E)-pyridinyl vinyl keto group was efficiently and specifically transferred to the 4-amino group of the opposing cytosine in RNA in the presence of NiCl2 with more than 200-fold accelerated rate compared with the previous system with the use of the diketo transfer group.Detailed mechanistic studies suggested that NiCl2 forms a bridging complex between the pyridinyl keto moiety and the N7 of the purine residue neighboring the cytosine residue of the RNA substrate to bring the groups in close proximity.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582 Japan, and CREST, Japan Science and Technology Agency, 4-1-8 Motomachi, Kawaguchi, Saitama 332-0012, Japan.

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Comparison of the transfer yields for RNA1. (A) The time course of the transfer yields. Closed squares: (E)-FT-ODN1 in the presence of NiCl2; open circles: (Z)-FT-ODN1 in the presence of NiCl2; closed diamonds: (E)-FT-ODN1 in the absence of NiCl2. (B) The transfer yields at 10 min. X represents the nucleotide opposite to the functionalized 6-thio-dG of FT-ODN1. a1 mM of Ethylenediaminetetraacetic acid (EDTA) was added. The reaction was performed at 37°C using 5 μM of RNA1 and 7.5 μM of FT-ODN1 in a buffer containing 50 mM HEPES (2-(4-(2-hydroxyethyl)piperazin-1-yl)ethane-1-sulfonic acid) and 100 mM NaCl at pH 7 in the presence or absence of NiCl2 (5 μM).
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Figure 3: Comparison of the transfer yields for RNA1. (A) The time course of the transfer yields. Closed squares: (E)-FT-ODN1 in the presence of NiCl2; open circles: (Z)-FT-ODN1 in the presence of NiCl2; closed diamonds: (E)-FT-ODN1 in the absence of NiCl2. (B) The transfer yields at 10 min. X represents the nucleotide opposite to the functionalized 6-thio-dG of FT-ODN1. a1 mM of Ethylenediaminetetraacetic acid (EDTA) was added. The reaction was performed at 37°C using 5 μM of RNA1 and 7.5 μM of FT-ODN1 in a buffer containing 50 mM HEPES (2-(4-(2-hydroxyethyl)piperazin-1-yl)ethane-1-sulfonic acid) and 100 mM NaCl at pH 7 in the presence or absence of NiCl2 (5 μM).

Mentions: Effect of metal cations on the transfer yield. (A) Comparison of the yields in the presence of 5 μM of MCl2. (B) Effect of the concentration of NiCl2. The transfer yields at 1 h are compared. The reaction conditions are the same as described in the footnote to Figure 3 except that the yield was measured after 60 min.


Remarkable acceleration of a DNA/RNA inter-strand functionality transfer reaction to modify a cytosine residue: the proximity effect via complexation with a metal cation.

Jitsuzaki D, Onizuka K, Nishimoto A, Oshiro I, Taniguchi Y, Sasaki S - Nucleic Acids Res. (2014)

Comparison of the transfer yields for RNA1. (A) The time course of the transfer yields. Closed squares: (E)-FT-ODN1 in the presence of NiCl2; open circles: (Z)-FT-ODN1 in the presence of NiCl2; closed diamonds: (E)-FT-ODN1 in the absence of NiCl2. (B) The transfer yields at 10 min. X represents the nucleotide opposite to the functionalized 6-thio-dG of FT-ODN1. a1 mM of Ethylenediaminetetraacetic acid (EDTA) was added. The reaction was performed at 37°C using 5 μM of RNA1 and 7.5 μM of FT-ODN1 in a buffer containing 50 mM HEPES (2-(4-(2-hydroxyethyl)piperazin-1-yl)ethane-1-sulfonic acid) and 100 mM NaCl at pH 7 in the presence or absence of NiCl2 (5 μM).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4117767&req=5

Figure 3: Comparison of the transfer yields for RNA1. (A) The time course of the transfer yields. Closed squares: (E)-FT-ODN1 in the presence of NiCl2; open circles: (Z)-FT-ODN1 in the presence of NiCl2; closed diamonds: (E)-FT-ODN1 in the absence of NiCl2. (B) The transfer yields at 10 min. X represents the nucleotide opposite to the functionalized 6-thio-dG of FT-ODN1. a1 mM of Ethylenediaminetetraacetic acid (EDTA) was added. The reaction was performed at 37°C using 5 μM of RNA1 and 7.5 μM of FT-ODN1 in a buffer containing 50 mM HEPES (2-(4-(2-hydroxyethyl)piperazin-1-yl)ethane-1-sulfonic acid) and 100 mM NaCl at pH 7 in the presence or absence of NiCl2 (5 μM).
Mentions: Effect of metal cations on the transfer yield. (A) Comparison of the yields in the presence of 5 μM of MCl2. (B) Effect of the concentration of NiCl2. The transfer yields at 1 h are compared. The reaction conditions are the same as described in the footnote to Figure 3 except that the yield was measured after 60 min.

Bottom Line: We have recently developed a new strategy for the in situ modification of RNA based on the functionality transfer reaction between an oligodeoxynucleotide probe and an RNA substrate. 2'-Deoxy-6-thioguanosine (6-thio-dG) was used as the platform to anchor the transfer group.It was demonstrated that the (E)-pyridinyl vinyl keto group was efficiently and specifically transferred to the 4-amino group of the opposing cytosine in RNA in the presence of NiCl2 with more than 200-fold accelerated rate compared with the previous system with the use of the diketo transfer group.Detailed mechanistic studies suggested that NiCl2 forms a bridging complex between the pyridinyl keto moiety and the N7 of the purine residue neighboring the cytosine residue of the RNA substrate to bring the groups in close proximity.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582 Japan, and CREST, Japan Science and Technology Agency, 4-1-8 Motomachi, Kawaguchi, Saitama 332-0012, Japan.

Show MeSH
Related in: MedlinePlus