A new approach for pyrophosphate bond formation starting from phosphoramidite derivatives by use of 6-trifluoromethyl-1-hydroxybenzotriazole-mediated O–N phosphoryl migration

Ohkubo, Akihiro; Aoki, Katsufumi; Seio, Kohji; Sekine, Mitsuo

doi:10.1016/j.tetlet.2003.11.110

Cited by 17 publications

(10 citation statements)

References 21 publications

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“…The chain elongation of modified oligonucleotides ( ODN 1 – 4 , as shown in Figure ) containing X 1 –X 4 was carried out in a DNA synthesizer using the standard phosphoramidite method. After the chain elongation, ODN 1 was deprotected and released from the CPG resin by treatment with 0.4 M NaOH in MeOH–H 2 O (4:1, v/v) at room temperature for 24 h. For the deprotection of the 2-cyanoethyl (CE) groups of the oligonucleotides containing X 2 and X 3 residue (with d -threoninol backbone), the CPG resins were treated with 10% DBU/pyridine–BSA (1:1, v/v) at room temperature for 3 h, , followed by treatment with a saturated aqueous ammonia solution at room temperature for 6 h to obtain ODN 2 – 3 . The ODN 4 was deprotected and released from the CPG resin by treatment with a saturated aqueous ammonia solution at room temperature for 2 h. ODN 1 – 4 were then characterized by MALDI-TOF mass spectrometry (see the Supporting Information).…”

Section: Resultsmentioning

confidence: 99%

“…The modified oligonucleotides were synthesized on a 1 μmol scale in an automated DNA/RNA synthesizer (ABI 392 DNA/RNA synthesizer) according to the standard phosphoramidite protocol of ultramild DNA synthesis (activator: 0.25 M 5-benzylthio-1 H -tetrazole in anhydrous MeCN, capping reagents: 5% phenoxyacetic anhydride in THF–pyridine and 10% 1-methylimidazole in THF, deblocking reagent: 3% trichloroacetic acid in CH 2 Cl 2 , and oxidizing reagent: 0.02 M I 2 in THF–pyridine–water). For the postsynthetic deprotection, ODN 1 was treated and cleaved (from CPG resin) with 0.4 M NaOH in MeOH–H 2 O (4:1, v/v) at room temperature for 24 h. ODN 2 – 3 were treated with 10% DBU/pyridine–BSA (1:1, v/v) at room temperature for 3 h, , followed by treatment with a saturated aqueous ammonia solution at room temperature for 6 h for their cleavage from CPG resin. ODN 4 was treated and cleaved (from CPG resin) with a saturated aqueous ammonia solution at room temperature for 6 h. The elution was removed by evaporation under reduced pressure, and the remaining was then purified on a Sep-pak C18 cartridge.…”

Section: Methodsmentioning

confidence: 99%

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pH-Dependent Switching of Base Pairs Using Artificial Nucleobases with Carboxyl Groups

et al. 2018

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In this study, we report the synthesis of modified oligonucleotides consisting of benzoic acid or isophthalic acid residues as new nucleobases. As evaluated by UV thermal denaturation analysis at different pH conditions (5.0, 6.0, 7.0, and 8.0), these modified oligonucleotides exhibited pH-dependent recognition of natural nucleobases and one is first found to be capable of base pair switching in response to a pH change. The isophthalic acid residue incorporated into the oligonucleotide on a d-threoninol backbone could preferentially bind with adenine but with guanine in response to a change in the pH conditions from pH 5 to pH 7 (or 8) without significant difference in duplex stability. These findings would be valuable for further developing pH-responsive DNA-based molecular devices.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

pH-Dependent Switching of Base Pairs Using Artificial Nucleobases with Carboxyl Groups

et al. 2018

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“…Accordingly, we developed pyrophosphorylating reagents with a phosphotriester structure, which is much more reactive than the phosphodiester structure of existing reagents. We developed the phosphotriester reagents by reference to a method for pyrophosphate bond formation reported by Sekine and coworkers ( 20 ). Pyrophosphorylating reagent 3 was prepared from phosphorylating reagent 1 and five equivalents of 1-hydroxy-6-(trifluoromethyl)benzotriazole (CF 3 -HOBt; Scheme 2 ).…”

Section: Resultsmentioning

confidence: 99%

Synthesis and biological activity of artificial mRNA prepared with novel phosphorylating reagents

Nagata

Hamasaki

Uetake

et al. 2010

Nucleic Acids Research

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Though medicines that target mRNA are under active investigation, there has been little or no effort to develop mRNA itself as a medicine. Here, we report the synthesis of a 130-nt mRNA sequence encoding a 33-amino-acid peptide that includes the sequence of glucagon-like peptide-1, a peptide that stimulates glucose-dependent insulin secretion from the pancreas. The synthesis method used, which had previously been developed in our laboratory, was based on the use of 2-cyanoethoxymethyl as the 2′-hydroxy protecting group. We also developed novel, highly reactive phosphotriester pyrophosphorylating reagents to pyrophosphorylate the 5′-end of the 130-mer RNA in preparation for capping. We completed the synthesis of the artificial mRNA by the enzymatic addition of a 5′-cap and a 3′-poly(A) tail to the pyrophosphorylated 130-mer and showed that the resulting mRNA supported protein synthesis in a cell-free system and in whole cells. As far as we know, this is the first time that mRNA has been prepared from a chemically synthesized RNA sequence. As well as providing a research tool for the intracellular expression of peptides, the technology described here may be used for the production of mRNA for medical applications.

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“…In the synthesis of 5′-short RNAs, we used the highly cross-linked polystyrene resin 8 1 that has a silyl-type linker, 9 which can be cleaved under neutral conditions, phosphoryl 10 and phosphityl reagents 2 – 4 (Figure 4 ). The synthesis of 5′-short RNA I is shown in Scheme 1 .…”

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confidence: 99%

Chemical Synthesis of U1 snRNA Derivatives

et al. 2013

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U1 snRNA is an interesting biological tool for splicing correction and regulation of gene expression. However, U1 snRNA has never been chemically synthesized. In this study, the first chemical synthesis of U1snRNA and its analogues was carried out. Moreover, it was found that the binding affinity of the modified U1 snRNA with an ethylene glycol linkage to snurportin 1 (nuclear import adaptor) was as high as that of the unmodified RNA.

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A new approach for pyrophosphate bond formation starting from phosphoramidite derivatives by use of 6-trifluoromethyl-1-hydroxybenzotriazole-mediated O–N phosphoryl migration

Cited by 17 publications

References 21 publications

pH-Dependent Switching of Base Pairs Using Artificial Nucleobases with Carboxyl Groups

pH-Dependent Switching of Base Pairs Using Artificial Nucleobases with Carboxyl Groups

Synthesis and biological activity of artificial mRNA prepared with novel phosphorylating reagents

Chemical Synthesis of U1 snRNA Derivatives

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