Reliable Chemical Synthesis of Oligoribonucleotides (RNA) with 2′-O-[(Triisopropylsilyl)oxy]methyl(2′-O-tom)-Protected Phosphoramidites

Pitsch, Stefan; Weiß, Patrick; Jenny, Luzi; Stutz, Alfred; Wu, Xiaolin

doi:10.1002/1522-2675(20011219)84:12<3773::aid-hlca3773>3.0.co;2-e

Cited by 221 publications

(200 citation statements)

References 17 publications

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“…Concerning the corresponding guanosine building blocks for the 2′-O-{[(triisopropylsilyl)oxy]methyl}(TOM) RNA approach, [14] we noted that the original paper reported high regioselectivity of the tomylation reaction (with a ratio of 12:1 in favor of the 2′-O over 3′-O isomer) [14] when the exocyclic N 2 amino group of the guanine moiety was protected with a single acyl group only. In analogy, we therefore decided to first cleave one of the two N 2 Boc groups of compound 1.…”

Section: Resultsmentioning

confidence: 99%

“…Then, tomylation of the 2′-hydroxyl group was conducted via the corresponding cyclic 2′,3′-O-di-tertbutylstannylidene derivative formed in situ in the presence of ethyldiisopropylamine and tBu 2 SnCl 2 in 1,2-dichloroethane at 50°C. Treatment with 1.3 equivalents of TOM-Cl [14] gave a mixture of 2′-O-and 3′-O-alkylated regioisomers in a favorable ratio of 4:1. The desired 2′-O-alkylated derivative 3b was easily isolated in pure form by chromatography on silica gel as the first-eluting isomer in 62% yield.…”

Section: Resultsmentioning

confidence: 99%

“…[10][11][12] The nucleoside building blocks require suitable protection of nucleobase amino and ribose 2′-hydroxy groups. Whereas the former are usually acyl protected, silyl protection (tBDMS or TOM) [13,14] of the latter are most widely applied, and orthoesters (ACE), [15] 2-cyanoethoxymethyl (CEM), [16] or thiocarbamates (TC) [17] also represent powerful alternatives. For all five approaches, the syntheses of the standard nucleoside phosphoramidites (A, C, G, U) have been very well optimized and are robust.…”

Section: Introductionmentioning

confidence: 99%

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An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

Jud

Micura

2017

Chemistry A European J

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We present an innovative O 6 -tert-butyl/N 2 -tert-butyloxycarbonyl protection concept for guanosine (G) phosphoramidites. This concept is advantageous for 2′-modified G building blocks because of very efficient synthetic access when compared with existing routes that usually employ O 6 -(4-nitrophenyl)ethyl/N 2 -acyl protection or that start from 2-aminoadenosine involving enzymatic transformation into guanosine later on in the synthetic path. The new phosphoramidites are fully compatible with 2′-O-tBDMS or TOM phosphoramidites in standard RNA solid-phase synthesis and deprotection, and provide excellent quality of tailored RNAs for the growing range of applications in RNA biophysics, biochemistry, and biology.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

Jud

Micura

2017

Chemistry A European J

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“…In all cases, the synthesis was at the 1.3 mM scale and the ribose 29-hydroxyl group of the building blocks was protected by the [(triisopropylsilyl)oxy]methyl (TOM) group (Pitsch et al 2001). All oligonucleotides were deprotected and cleaved from solid support using the following conditions: (1) CH 3 NH 2 in ethanol (8 M, 700 mL) and CH 3 NH 2 in water (41%, 700 mL) for 6 h; (2) TBAFÁ3H 2 O in THF (1 M, 950 mL) for 12 h; and (3) neutralization with triethylammonium acetate buffer in water (1 M, 950 mL).…”

Section: Oligonucleotide Synthesis and Purificationmentioning

confidence: 99%

Effects of N²,N²-dimethylguanosine on RNA structure and stability: Crystal structure of an RNA duplex with tandem m²₂G:A pairs

Pallan¹,

Kreutz²,

Bosio³

et al. 2008

RNA

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Methylation of the exocyclic amino group of guanine is a relatively common modification in rRNA and tRNA. ,N 2 -dimethyl modification is its role in controlling the pairing modes between G and A. We have determined the crystal structure of a 13-mer RNA duplex with central tandem m 2 2 G:A pairs. In the structure both pairs adopt an imino-hydrogen bonded, pseudo-Watson-Crick conformation. Thus, the sheared conformation frequently seen in tandem G:A pairs is avoided due to a potential steric clash between an N 2 -methyl group and the major groove edge of A. Additionally, for a series of G:A containing self-complementary RNAs we investigated how methylation affects competitive hairpin versus duplex formation based on UV melting profile analysis.

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“…In 1998, Pitsch and coworkers developed a novel 2′-O-protecting group, 2′-O-TOM ( Figure 3, panel b), that gives high coupling yields (>99%) due to reduced steric hindrance compared to TBDMS (25,26). With this strategy, long (up to 80 nt) RNAs with site-specific modifications can be generated.…”

mentioning

confidence: 99%

Combined Approaches to Site-Specific Modification of RNA

2008

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Both natural and unnatural modifications in RNA are of interest to biologists and chemists. More than 100 different analogs of the four standard RNA nucleosides have been identified in nature. Unnatural modifications are useful for structure and mechanistic studies of RNA. This Review highlights chemical, enzymatic, and combined (semisynthesis) approaches to generate sitespecifically modified RNAs. The availability of these methods for site-specific modifications of RNAs of all sizes is important in order to study the relationships between RNA chemical composition, structure, and function.RNAs undergo specific post-transcriptional modification by a wide variety of enzymes and ribonucleoprotein complexes (1,2). More than 100 different modifications of the four standard RNA nucleosides, adenosine, cytidine, guanosine, and uridine, have been identified (3). Examples from each of the four major categories of base isomerization, base modification, sugar modification, and hypermodification are shown in Figure 1, panel a. Similarly, a large number of unnatural modifications have been synthesized and used for structure and mechanistic studies of RNA (4-6). A few examples are shown in Figure 1, panel b (7-10). Natural modified nucleotides are often found in the functionally important regions of RNA, such as the peptidyl transferase center of ribosomal RNA (rRNA), the anticodon loop of transfer RNAs, or the branch site of spliceosomal . Several modifications, such as pseudouridine (14), have been known for almost 50 years. Their locations in natural RNAs can, in some cases, be highly conserved; yet, our understanding of their biological roles is still incomplete (15). Through a combination of biological, chemical, and biophysical approaches, much can be learned regarding the roles of modified nucleotides. Similarly, the incorporation of unnatural modifications may be useful in order to study the biological roles and functions of RNA (5).In a number of cases, the enzymes or small nucleolar RNAs (snoRNAs) that are responsible for site-specific RNA modification have been identified (2). Knock-out studies of the individual modifying enzymes or snoRNAs then allow the function of the modification to be deduced. In cases where the enzymes are not known, or the modification is not a natural one, alternative approaches must be considered. Over the past several decades, several chemical methods have been developed to study the effects of modifications in small RNA model systems. These studies have led to newer, more powerful approaches that allow site-specific modification of large RNAs, reconstitution into biological systems, and studies of RNA structure and function. Chemical SynthesisChemical synthesis of RNA allows for site-selective incorporation of modified nucleotides. Five decades ago, Michelson and coworkers synthesized a thymidylate dinucleotide by the *Corresponding author csc@chem.wayne.edu. Figure 2): i) coupling at the 5′ site with a protected phosphoramidite, ii) capping of the unreacted 5′-hydroxyl groups, ...

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Reliable Chemical Synthesis of Oligoribonucleotides (RNA) with 2′-O-[(Triisopropylsilyl)oxy]methyl(2′-O-tom)-Protected Phosphoramidites

Cited by 221 publications

References 17 publications

An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

Effects of N²,N²-dimethylguanosine on RNA structure and stability: Crystal structure of an RNA duplex with tandem m²₂G:A pairs

Combined Approaches to Site-Specific Modification of RNA

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Reliable Chemical Synthesis of Oligoribonucleotides (RNA) with 2′-O-[(Triisopropylsilyl)oxy]methyl(2′-O-tom)-Protected Phosphoramidites

Cited by 221 publications

References 17 publications

An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

An Unconventional Acid‐Labile Nucleobase Protection Concept for Guanosine Phosphoramidites in RNA Solid‐Phase Synthesis

Effects of N2,N2-dimethylguanosine on RNA structure and stability: Crystal structure of an RNA duplex with tandem m22G:A pairs

Combined Approaches to Site-Specific Modification of RNA

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Effects of N²,N²-dimethylguanosine on RNA structure and stability: Crystal structure of an RNA duplex with tandem m²₂G:A pairs