Novel method of synthesis of 5''-phosphate 2'-O-ribosyl-ribonucleosides and their 3'-phosphoramidites.

Marcin K Chmielewski,Wojciech T Markiewicz

doi:10.3390/molecules181214780

Marcin K Chmielewski, Wojciech T Markiewicz

Open Access

https://doi.org/10.3390/molecules181214780

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Abstract

Synthesis of 5''-phosphate 2'-O-ribosylribonucleosides [Nr(p)] of four common ribonucleosides, and 3'-phosphoramidites of 5''-phosphate 2'-O-ribosyladenosine and 2'-O-ribosylguanosine using the H-phosphonate chemistry is described. An additional ring protected by benzoyl groups was incorporated into the main ribosyl ring in the reaction with 1-O-acetyl-2,3,5-tri-O-benzoyl-β-d-ribofuranose in the presence of SnCl4. The obtained 2'-O-ribosylribonucleosides (Nr) were applied in the subsequent transformations with selective deprotection. Ethanolamine was applied as a very convenient reagent for selective removal of benzoyl groups. Additionally, the tetraisopropyldisiloxane-1,3-diyl (TIPDSi) group was found to be stable under these deprotection conditions. Thus, the selectively deprotected 5''-hydroxyl group of Nr was transformed into an H-phosphonate monoester which was found to be stable under the following conditions: the removal of the TIPDSi group with triethylammonium fluoride and the dimethoxytritylation of the 5''-hydroxyl function. The 5''-H-phosphonate of Nr precursors was easily transformed to the corresponding dicyanoethyl 5''-O-phosphotriesters before phosphitylation, which gave 3'-phosphoramidite units of Nr(p) in high yield. The derived phosphoramidite units were used in an automated oligonucleotide synthesizer to produce dimer Ar(p)T via the phosphoramidite approach. The obtained products were fully deprotected under standard deprotection conditions giving dimers with a 5''-phosphate monoester function. Application of an alkaline phosphatase to prove the presence of an additional phosphate group was described.

Highlights

The 2'-O-ribosylribonucleosides, which are the most common natural representatives of disaccharide nucleosides, were found as minor nucleosides in tRNA with a key function in protein biosynthesis
The derived phosphoramidite units were used in an automated oligonucleotide synthesizer to produce dimer Ar(p)T via the phosphoramidite approach
Chemical synthesis of 5''-phosphorylated disaccharide nucleosides requires choosing a proper protecting system in order to handle the phosphate functional groups that are to be introduced into different positions and handled differently throughout the synthesis of monomers and their later introduction into oligonucleotides

Summary

Introduction

The 2'-O-ribosylribonucleosides, which are the most common natural representatives of disaccharide nucleosides, were found as minor nucleosides in tRNA with a key function in protein biosynthesis. The presence of 2'-O-ribosyl modified nucleoside in tRNA determines the functions of methionine tRNA as initiation tRNA [4] Discovery of this important modification in natural products started the investigation process aimed at their possible applications. 2'-O-β-D-ribofuranosyl-(1''-2')-guanosine-5''-O-phosphate [15] was developed, the yields were low These compounds were applied in chemical synthesis of nucleic acids and incorporated in RNA sequences with other minor components of tRNA [16]. This modification has no effect on thermal stability of the duplex, and the extra ribose moiety seems to be stabilized by bridged H-bonds [17]. Enzymatic incorporation of 2'-O-ribosylribonucleoside residue into oligonucleotides was investigated [18]

Results and Discussion

General Methods

Conclusions