Synthesis of Capped RNA using a DMT Group as a Purification Handle

Veliath, Elizabeth; Gaffney, Barbara L.; Jones, R. A. C.

doi:10.1080/15257770.2013.864417

Cited by 8 publications

(8 citation statements)

References 21 publications

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“…Short m 7 GpppRNAs, m 3 GpppRNAs, and NAD-RNAs have been synthesized by chemical reaction of RNA-5′ phosphates with appropriate imidazole-activated nucleotides under aqueous conditions [ 109 – 112 ], but these reactions are only moderately efficient and require time-consuming chromatographic and/or enzymatic work-ups to separate capped and uncapped RNAs. An improvement in solution synthesis of capped oligomers has been proposed by using a 4,4′-dimethoxytrityl (DMT) group as a lipophilic purification handle, which facilitates separation of capped RNAs from uncapped RNAs by RP HPLC [ 113 ]. Solid-phase synthesis of capped oligonucleotides has been attempted as well, but found to be challenging due to incompatibility of the m 7 G moiety with standard oligonucleotide de-immobilization and de-protection protocols [ 114 – 118 ].…”

Section: Chemically and Enzymatically Labeled Rna Capsmentioning

confidence: 99%

Applications of Phosphate Modification and Labeling to Study (m)RNA Caps

et al. 2017

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The cap is a natural modification present at the 5′ ends of eukaryotic messenger RNA (mRNA), which because of its unique structural features, mediates essential biological functions during the process of gene expression. The core structural feature of the mRNA cap is an N7-methylguanosine moiety linked by a 5′–5′ triphosphate chain to the first transcribed nucleotide. Interestingly, other RNA 5′ end modifications structurally and functionally resembling the m7G cap have been discovered in different RNA types and in different organisms. All these structures contain the ‘inverted’ 5′–5′ oligophosphate bridge, which is necessary for interaction with specific proteins and also serves as a cleavage site for phosphohydrolases regulating RNA turnover. Therefore, cap analogs containing oligophosphate chain modifications or carrying spectroscopic labels attached to phosphate moieties serve as attractive molecular tools for studies on RNA metabolism and modification of natural RNA properties. Here, we review chemical, enzymatic, and chemoenzymatic approaches that enable preparation of modified cap structures and RNAs carrying such structures, with emphasis on phosphate-modified mRNA cap analogs and their potential applications.

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Section: Chemically and Enzymatically Labeled Rna Capsmentioning

confidence: 99%

Applications of Phosphate Modification and Labeling to Study (m)RNA Caps

et al. 2017

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“…18 The synthesis of GMPS has been reported previously but oen involving tedious and unconventional procedures. [26][27][28][29][30][31] We synthesized 6 from 1 by rst protecting the 2 0 and 3 0 hydroxyl groups with acetone through ketal formation to afford 2 with an excellent yield (98%, Scheme 2). 32 We initially attempted to directly synthesize 6 by thiophosphorylating 2, producing an ammonium salt in the presence of NH 4 OH, and nally, removing the acetone protection group to attain the desired 6 (the "R"-shaped arrow in Scheme 2).…”

Section: Resultsmentioning

confidence: 99%

Corroboration of Zn(ii)–Mg(ii)-tertiary structure interplays essential for the optimal catalysis of a phosphorothiolate thiolesterase ribozyme

Wang

Chen

et al. 2018

RSC Adv.

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The TW17 ribozyme, a catalytic RNA selected from a pool of artificial RNA, is specific for the Zn 2+dependent hydrolysis of a phosphorothiolate thiolester bond. Here, we describe the organic synthesis of both guanosine a-thio-monophosphate and the substrates required for selecting and characterizing the TW17 ribozyme, and for deciphering the catalytic mechanism of the ribozyme. By successively substituting the substrate originally conjugated to the RNA pool with structurally modified substrates, we demonstrated that the TW17 ribozyme specifically catalyzes phosphorothiolate thiolester hydrolysis.Metal titration studies of TW17 ribozyme catalysis in the presence of Zn 2+ alone, Zn 2+ and Mg 2+ , and Zn 2+ and [Co(NH 3 ) 6 ] 3+ supported our findings that Zn 2+ is absolutely required for ribozyme catalysis, and indicated that optimal ribozyme catalysis involves the presence of outer-sphere and one inner-sphere Mg 2+ . A survey of the TW17 ribozyme activity at various pHs revealed that the activity of the ribozyme critically depends on the alkaline conditions. Moreover, a GNRA tetraloop-containing ribozyme constructed with active catalysis in trans provided catalysis and multiple substrate turnover efficiencies significantly higher than ribozymes lacking a GNRA tetraloop. This research supports the essential roles of Zn 2+ , Mg 2+ , and a GNRA tetraloop in modulating the TW17 ribozyme structure for optimal ribozyme catalysis, leading also to the formulation of a proposed reaction mechanism for TW17 ribozyme catalysis. Taiwan † Electronic supplementary information (ESI) available: Full details of supporting gures referenced in the text, NMR and MS spectra for some known and all the new compounds (2, 3, 6, 8b, 9a, 9b, 10a, 10b, 15b, 16b, 17, 18a, and 18b). See Scheme 2 Synthesis of the ammonium salt of guanosine a-thio-monophosphate (6). Three routes for the synthesis of 6 were explored in this study. Only the route sequentially synthesizing 4 and 5 from 2 could produce 6 with a high yield (see text). This journal isScheme 3 Synthesis of substrates and substrate-RNA conjugates. (A) Synthesis of the substrates required for the in vitro selection of the TW17 ribozyme and the determination of the substrate specificity of the TW17 ribozyme. (B) Preparation of the substrate-RNA conjugates for in vitro evolution and selection of the TW17 ribozyme. 32778 | RSC Adv., 2018, 8, 32775-32793This journal is Scheme 4 Synthesis of the substrates utilized in studying the Zn 2+ -dependent hydrolytic mechanism of the TW17 ribozyme. This journal isScheme 5 The proposed mechanism of optimal TW17 ribozyme catalysis.32786 | RSC Adv., 2018, 8, 32775-32793This journal is

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“…It is also worth noting that the addition of excess of base (10 equiv) is required to keep the reaction medium basic and avoid deprotection of the DMT group. 43 With the reaction parameters having been optimized, studies on the scope of the process were undertaken for the synthesis of several nucleoside-based amides by incorporating a variety of different amines. Various benzyl, heteroaryl, acyclic and cyclic alkyl amines were utilized and were proficiently transformed into the desired products 3a-v in good to excellent yields (Scheme 2).…”

Section: Paper Syn Thesismentioning

confidence: 99%

Pd/PTABS: An Efficient Catalytic System for the Aminocarbonylation of a Sugar-Protected Nucleoside

et al. 2019

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A highly efficient and mild protocol for the aminocarbonylation of a nucleoside is developed by employing palladium/(1,3,5-triaza-7-phosphaadamantan-1-ium-1-yl)butane-1-sulfonate (Pd/PTABS) as the catalytic system. The developed aminocarbonylation methodology employs CO gas at a relatively low temperature of 60 °C and is suitable for a wide range of amines, including (heteroaryl)benzylic, aliphatic acyclic, alicyclic and secondary amines. This protocol is also utilized for the synthesis of a sangivamycin precursor by carrying out the Pd-catalyzed amination and aminocarbonylation simultaneously. The utility of this protocol is further demonstrated by the synthesis of the drugs moclobemide and nikethamide.

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Synthesis of Capped RNA using a DMT Group as a Purification Handle

Cited by 8 publications

References 21 publications

Applications of Phosphate Modification and Labeling to Study (m)RNA Caps

Applications of Phosphate Modification and Labeling to Study (m)RNA Caps

Corroboration of Zn(ii)–Mg(ii)-tertiary structure interplays essential for the optimal catalysis of a phosphorothiolate thiolesterase ribozyme

Pd/PTABS: An Efficient Catalytic System for the Aminocarbonylation of a Sugar-Protected Nucleoside

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