Hidehito Urata - Cochemist

Syntheses of prodrug-type phosphotriester oligonucleotides responsive to intracellular reducing environment for improvement of cell membrane permeability and nuclease resistance

Co-reporter:Junsuke Hayashi, Yusuke Samezawa, Yosuke Ochi, Shun-ichi Wada, Hidehito Urata

Bioorganic & Medicinal Chemistry Letters 2017 Volume 27, Issue 14(Issue 14) pp:

Publication Date(Web):15 July 2017

DOI:10.1016/j.bmcl.2017.05.031

We synthesized prodrug-type phosphotriester (PTE) oligonucleotides containing the six-membered cyclic disulfide moiety by using phosphoramidite chemistry. Prodrug-type oligonucleotides named “Reducing-Environment-Dependent Uncatalyzed Chemical Transforming (REDUCT) PTE oligonucleotides” were converted into natural oligonucleotides under cytosol-mimetic reductive condition. Furthermore, the REDUCT PTE oligonucleotides were robust to nuclease digestion and exhibited good cell membrane permeability.Download high-res image (66KB)Download full-size image

Gene silencing by 2′-O-methyldithiomethyl-modified siRNA, a prodrug-type siRNA responsive to reducing environment

Co-reporter:Yosuke Ochi, Mieko Imai, Osamu Nakagawa, Junsuke Hayashi, Shun-ichi Wada, Hidehito Urata

Bioorganic & Medicinal Chemistry Letters 2016 Volume 26(Issue 3) pp:845-848

Publication Date(Web):1 February 2016

DOI:10.1016/j.bmcl.2015.12.074

RNAs bearing various 2′-modifications have been synthesized in an effort to improve nuclease resistance. However, the gene silencing activity of small interfering RNAs (siRNAs) has been decreased or sometimes completely suppressed by the chemical modifications. We previously developed a post-synthetic approach for the synthesis of 2′-O-methyldithiomethyl-modified RNA, which can be converted into unmodified RNA under reducing conditions, and named it Reducing-Environment-Dependent Uncatalyzed Chemical Transforming RNA (REDUCT RNA). Here, the gene silencing activity of REDUCT siRNA bearing 2′-O-methyldithiomethyl groups was evaluated. REDUCT siRNA showed more effective gene silencing than unmodified siRNA regardless of the modification site. This result suggests that REDUCT siRNA is converted into unmodified siRNA inside cells as a prodrug-type siRNA.

Regulated Incorporation of Two Different Metal Ions into Programmed Sites in a Duplex by DNA Polymerase Catalyzed Primer Extension

Co-reporter:Tatsuya Funai;Junko Nakamura;Yuki Miyazaki;Risa Kiriu;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Akira Ono; Hidehito Urata

Angewandte Chemie 2014 Volume 126( Issue 26) pp:6742-6745

Publication Date(Web):

DOI:10.1002/ange.201311235

Abstract

Metal-mediated base pairs formed by the coordination of metal ions to natural or artificial bases impart unique chemical and physical properties to nucleic acids and have attracted considerable interest in the field of nanodevices. Ag^I ions were found to mediate DNA polymerase catalyzed primer extension through the formation of a C–Ag^I–T base pair, as well as the previously reported C–Ag^I–A base pair. The comparative susceptibility of dNTPs to Ag^I-mediated enzymatic incorporation into the site opposite cytosine in the template was shown to be dATP>dTTP≫dCTP. Furthermore, two kinds of metal ions, Ag^I and Hg^II, selectively mediate the incorporation of thymidine 5′-triphosphate into sites opposite cytosine and thymine in the template, respectively. In other words, the regulated incorporation of different metal ions into programmed sites in the duplex by DNA polymerase was successfully achieved.

Regulated Incorporation of Two Different Metal Ions into Programmed Sites in a Duplex by DNA Polymerase Catalyzed Primer Extension

Co-reporter:Tatsuya Funai;Junko Nakamura;Yuki Miyazaki;Risa Kiriu;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Akira Ono; Hidehito Urata

Angewandte Chemie International Edition 2014 Volume 53( Issue 26) pp:6624-6627

Publication Date(Web):

DOI:10.1002/anie.201311235

Abstract

Metal-mediated base pairs formed by the coordination of metal ions to natural or artificial bases impart unique chemical and physical properties to nucleic acids and have attracted considerable interest in the field of nanodevices. Ag^I ions were found to mediate DNA polymerase catalyzed primer extension through the formation of a C–Ag^I–T base pair, as well as the previously reported C–Ag^I–A base pair. The comparative susceptibility of dNTPs to Ag^I-mediated enzymatic incorporation into the site opposite cytosine in the template was shown to be dATP>dTTP≫dCTP. Furthermore, two kinds of metal ions, Ag^I and Hg^II, selectively mediate the incorporation of thymidine 5′-triphosphate into sites opposite cytosine and thymine in the template, respectively. In other words, the regulated incorporation of different metal ions into programmed sites in the duplex by DNA polymerase was successfully achieved.

Rcktitelbild: Regulated Incorporation of Two Different Metal Ions into Programmed Sites in a Duplex by DNA Polymerase Catalyzed Primer Extension (Angew. Chem. 26/2014)

Co-reporter:Tatsuya Funai;Junko Nakamura;Yuki Miyazaki;Risa Kiriu;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Akira Ono; Hidehito Urata

Angewandte Chemie 2014 Volume 126( Issue 26) pp:

Publication Date(Web):

DOI:10.1002/ange.201401152

Back Cover: Regulated Incorporation of Two Different Metal Ions into Programmed Sites in a Duplex by DNA Polymerase Catalyzed Primer Extension (Angew. Chem. Int. Ed. 26/2014)

Co-reporter:Tatsuya Funai;Junko Nakamura;Yuki Miyazaki;Risa Kiriu;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Akira Ono; Hidehito Urata

Angewandte Chemie International Edition 2014 Volume 53( Issue 26) pp:

Publication Date(Web):

DOI:10.1002/anie.201401152

A post-synthetic approach for the synthesis of 2′-O-methyldithiomethyl-modified oligonucleotides responsive to a reducing environment

Co-reporter:Yosuke Ochi, Osamu Nakagawa, Katsunori Sakaguchi, Shun-ichi Wada and Hidehito Urata

Chemical Communications 2013 vol. 49(Issue 69) pp:7620-7622

Publication Date(Web):27 Jun 2013

DOI:10.1039/C3CC43725F

Based on a novel concept, a Reducing-Environment-Dependent Uncatalyzed Chemical Transforming RNA, “REDUCT RNA”, we established a post-synthetic approach for the synthesis of 2′-O-methyldithiomethyl-modified oligonucleotides from 2′-O-(2,4,6-trimethoxybenzylthiomethyl)-oligonucleotides by treatment with dimethyl(methylthio)sulfonium tetrafluoroborate. 2′-O-methyldithiomethyl oligonucleotides were easily converted into 2′-hydroxy oligonucleotides under reducing conditions, such as those found in the intracellular environment.

Thermal stability of oligodeoxynucleotide duplexes containing l-deoxynucleotide at termini

Co-reporter:Hidehito Urata, Shuji Ogawa, Shun-ichi Wada

Bioorganic & Medicinal Chemistry Letters 2013 Volume 23(Issue 10) pp:2909-2911

Publication Date(Web):15 May 2013

DOI:10.1016/j.bmcl.2013.03.058

The effects of substituting l-deoxynucleotide for d-deoxynucleotide at duplex termini were evaluated and the terminal substitutions were found to show much less effects on duplex destabilization and to show a similar tendency in base pairing selectivity, compared with internal chiral substitutions.Terminal chiral modifications of oligodeoxynucleotide duplexes show much less effects on duplex destabilization and to show a similar tendency in base pairing selectivity, compared with internal chiral modifications.

Base recognition by L-nucleotides in heterochiral DNA

Co-reporter:Shuji Ogawa, Shun-ichi Wada and Hidehito Urata

RSC Advances 2012 vol. 2(Issue 6) pp:2274-2275

Publication Date(Web):06 Feb 2012

DOI:10.1039/C2RA01013E

Thermal stabilities of homo- and heterochiral DNA duplexes with and without a base pair mismatch at the chiral modification site were evaluated. The results indicated that the L-nucleotide residue in heterochiral duplexes retains its selectivity toward the complementary D-nucleotide residue.

AgI Ion Mediated Formation of a CA Mispair by DNA Polymerases

Co-reporter:Tatsuya Funai;Yuki Miyazaki;Megumi Aotani;Eriko Yamaguchi;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Hidetaka Torigoe; Akira Ono; Hidehito Urata

Angewandte Chemie International Edition 2012 Volume 51( Issue 26) pp:6464-6466

Publication Date(Web):

DOI:10.1002/anie.201109191

AgI Ion Mediated Formation of a CA Mispair by DNA Polymerases

Co-reporter:Tatsuya Funai;Yuki Miyazaki;Megumi Aotani;Eriko Yamaguchi;Dr. Osamu Nakagawa;Dr. Shun-ichi Wada; Hidetaka Torigoe; Akira Ono; Hidehito Urata

Angewandte Chemie 2012 Volume 124( Issue 26) pp:6570-6572

Publication Date(Web):

DOI:10.1002/ange.201109191

Pyrimidine–pyrimidine base pairs stabilized by silver(I) ions

Co-reporter:Hidehito Urata, Eriko Yamaguchi, Yasunari Nakamura and Shun-ichi Wada

Chemical Communications 2011 vol. 47(Issue 3) pp:941-943

Publication Date(Web):15 Nov 2010

DOI:10.1039/C0CC04091F

In the presence of AgI ions, the C–T and m5iC (5-methylisocytosine)–T base pairs showed comparable stability to the C–AgI–C base pair, and the m5iC–C base pair was highly stabilized by the synergetic effect of AgI coordination and possible hydrogen bonding.

Facile and efficient approach for the synthesis of N2-dimethylaminomethylene-2′-O-methylguanosine

Co-reporter:Tsuyoshi Mukobata, Yosuke Ochi, Yuji Ito, Shun-ichi Wada, Hidehito Urata

Bioorganic & Medicinal Chemistry Letters 2010 Volume 20(Issue 1) pp:129-131

Publication Date(Web):1 January 2010

DOI:10.1016/j.bmcl.2009.11.016

A convenient method for the synthesis of N2-dimethylaminomethylene-2′-O-methylguanosine (1), which is a useful intermediate for oligonucleotide construction, was developed. We chose the di-tert-butylsilyl group and the triisopropylbenzenesulfonyl group as sugar and base protecting groups, respectively. These protecting groups were stable during the 2′-O-methylation step with MeI and NaH. Our six-step synthesis of 1 is easy to perform using commercially available reagents, and requires only three chromatographic purifications. Compound 1 was obtained in 56% yield from guanosine.

Incorporation of Thymine Nucleotides by DNA Polymerases through THgIIT Base Pairing

Co-reporter:Hidehito Urata ;Eriko Yamaguchi;Tatsuya Funai;Yuriko Matsumura ;Shun-ichi Wada Dr.

Angewandte Chemie 2010 Volume 122( Issue 37) pp:6666-6669

Publication Date(Web):

DOI:10.1002/ange.201002142

Incorporation of Thymine Nucleotides by DNA Polymerases through THgIIT Base Pairing

Co-reporter:Hidehito Urata ;Eriko Yamaguchi;Tatsuya Funai;Yuriko Matsumura ;Shun-ichi Wada Dr.

Angewandte Chemie International Edition 2010 Volume 49( Issue 37) pp:6516-6519

Publication Date(Web):

DOI:10.1002/anie.201002142

Kinetic analysis of hydrolytic reaction of homo- and heterochiral adenylyl(3′–5′)adenosine isomers: breaking homochirality reduces hydrolytic stability of RNA

Co-reporter:Hidehito Urata, Rie Sasaki, Hiroyo Morita, Marina Kusumoto, Yoko Ogawa, Kozue Mitsuda and Masao Akagi

Chemical Communications 2005 (Issue 20) pp:2578-2580

Publication Date(Web):18 Mar 2005

DOI:10.1039/B500673B

The hydrolytic stability of the diastereomeric isomers of ApA was compared and the results show that heterochiral ApAs are more rapidly hydrolyzed than homochiral ApAs at low temperatures, suggesting that hydrolytic selection in cold environments in conjunction with selective polymerization may have been effective in enriching the homochirality of RNA.

Synthesis and structural characterization of diastereomeric isomers of RNA trimer adenylyl(3′-5′)adenylyl(3′-5′)adenosine

Co-reporter:Hidehito Urata, Hisafumi Hara, Yoshihiro Hirata, Norihiko Ohmoto, Masao Akagi

Tetrahedron: Asymmetry 2005 Volume 16(Issue 17) pp:2908-2917

Publication Date(Web):5 September 2005

DOI:10.1016/j.tetasy.2005.07.026

We have synthesized the diastereomeric isomers of adenylyl(3′-5′)adenylyl(3′-5′)adenosine (ApApA), and investigated their helical structures and hybridization properties with d-poly(U) by circular dichroism (CD) and UV melting experiments. The configuration of the 5′-end residue of ApApA has little effect on the helical structure. The configuration of the 3′-end residue has little effect on the stability of the triplex, and the chiral modifications cause complicated effects on the helical structure of ApApA and its triplex-forming ability with d-poly(U), depending on the site of the modification.

Synthesis and hybridization properties of L-oligodeoxynucleotide analogues fixed in a low anti glycosyl conformation

Co-reporter:Hidehito Urata, Hidetaka Miyagoshi, Tetsuya Kumashiro, Takashi Yumoto, Keiji Mori, Keiko Shoji, Keigo Gohda and Masao Akagi

Organic & Biomolecular Chemistry 2004 vol. 2(Issue 2) pp:183-189

Publication Date(Web):10 Dec 2003

DOI:10.1039/B312276J

We have synthesized L-type enantiomers (cU and cA) of nucleoside analogues, whose glycosyl bonds are fixed in a low anti conformation (ap glycosyl conformation, χ ≈ 180°), and incorporated them into oligonucleotides to evaluate the hybridization ability with natural DNA and RNA sequences. Although the incorporation of the modified nucleosides into oligonucleotides decreased the hybridization ability with unmodified complementary DNA sequences, the fully-substituted 12mers (cU12 and cA12) still retained the hybridization ability with the complementary unmodified DNA 12mers, regardless of their unnatural L-chirality. In contrast, cU12 and cA12 showed different hybridization behavior with complementary unmodified RNA 12mers. cU12 forms a more stable duplex with rA12 than the corresponding natural 12mer (dT12), whereas cA12 cannot hybridize with rU12. Based on the model structure of cU12–rA12, we discuss these experimental results.

Helical structure of heterochiral RNA dimers: helical sense of ApA is determined by chirality of 3′-end residue

Co-reporter:Hidehito Urata, Makiko Go, Norihiko Ohmoto, Katsuhiko Minoura and Masao Akagi

Chemical Communications 2002 (Issue 5) pp:544-545

Publication Date(Web):14 Feb 2002

DOI:10.1039/B111312G

Heterochiral ApAs (ALpAL and ALpAL) have been synthesized and investigation of their helical structures by means of spectroscopic techniques indicates that the chirality of the 3′-end residue is the primary factor for determining the helical sense of ApA.

Pyrimidine–pyrimidine base pairs stabilized by silver(I) ions

Co-reporter:Hidehito Urata, Eriko Yamaguchi, Yasunari Nakamura and Shun-ichi Wada

Chemical Communications 2011 - vol. 47(Issue 3) pp:NaN943-943

Publication Date(Web):2010/11/15

DOI:10.1039/C0CC04091F

In the presence of AgI ions, the C–T and m5iC (5-methylisocytosine)–T base pairs showed comparable stability to the C–AgI–C base pair, and the m5iC–C base pair was highly stabilized by the synergetic effect of AgI coordination and possible hydrogen bonding.

A post-synthetic approach for the synthesis of 2′-O-methyldithiomethyl-modified oligonucleotides responsive to a reducing environment

Co-reporter:Yosuke Ochi, Osamu Nakagawa, Katsunori Sakaguchi, Shun-ichi Wada and Hidehito Urata

Chemical Communications 2013 - vol. 49(Issue 69) pp:NaN7622-7622

Publication Date(Web):2013/06/27

DOI:10.1039/C3CC43725F

Based on a novel concept, a Reducing-Environment-Dependent Uncatalyzed Chemical Transforming RNA, “REDUCT RNA”, we established a post-synthetic approach for the synthesis of 2′-O-methyldithiomethyl-modified oligonucleotides from 2′-O-(2,4,6-trimethoxybenzylthiomethyl)-oligonucleotides by treatment with dimethyl(methylthio)sulfonium tetrafluoroborate. 2′-O-methyldithiomethyl oligonucleotides were easily converted into 2′-hydroxy oligonucleotides under reducing conditions, such as those found in the intracellular environment.