Microwave‐Assisted Phosphitylation of DNA and RNA Nucleosides and Their Analogs

Efthymiou, Tim C.; Krishnamurthy, Ramanarayanan

doi:10.1002/0471142700.nc0219s60

Cited by 4 publications

(3 citation statements)

References 18 publications

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“…The silyl acetal was selectively removed using hydrogen fluoride ( 4 ), which was then 5′- O -4,4′-dimethoxytrityl (DMT) protected ( 5 ) . The 3′- O -phosphoramidite was installed via a microwave-assisted synthesis protocol developed by Krishnamurthy and co-workers to afford BFU-phosphoramidite 6 . , The overall yield from commercial starting materials was 35–45%, as compared to the ∼28% overall yield reported for an alternate method reported after the start of this work . Solid-phase synthesis and purification are described in the Supporting Information, S3.…”

Section: Resultsmentioning

confidence: 99%

Small Molecule-Based Pattern Recognition To Classify RNA Structure

et al. 2016

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Three-dimensional RNA structures are notoriously difficult to determine, and the link between secondary structure and RNA conformation is only beginning to be understood. These challenges have hindered the identification of guiding principles for small molecule:RNA recognition. We herein demonstrate that the strong and differential binding ability of aminoglycosides to RNA structures can be used to classify five canonical RNA secondary structure motifs through principal component analysis (PCA). In these analyses, the aminoglycosides act as receptors, while RNA structures labeled with a benzofuranyluridine fluorophore act as analytes. Complete (100%) predictive ability for this RNA training set was achieved by incorporating two exhaustively guanidinylated aminoglycosides into the receptor library. The PCA was then externally validated using biologically relevant RNA constructs. In bulge-stem-loop constructs of HIV-1 transactivation response element (TAR) RNA, we achieved nucleotide-specific classification of two independent secondary structure motifs. Furthermore, examination of cheminformatic parameters and PCA loading factors revealed trends in aminoglycoside:RNA recognition, including the importance of shape-based discrimination, and suggested the potential for size and sequence discrimination within RNA structural motifs. These studies present a new approach to classifying RNA structure and provide direct evidence that RNA topology, in addition to sequence, is critical for the molecular recognition of RNA.

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Section: Resultsmentioning

confidence: 99%

Small Molecule-Based Pattern Recognition To Classify RNA Structure

et al. 2016

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“…In order to shorten the duration of the reaction we applied microwave‐heating. This idea was inspired by our previous use of microwaves for the preparation of phosphoramidites by phosphitylation of RNA and DNA nucleosides (Efthymiou & Krishnamurthy, 2015; Meher et al., 2014) and synthesis of cyclic‐phospholipids by combining microwaves and BDMDAP (Ortuno et al., 2022).…”

Section: Commentarymentioning

confidence: 99%

Microwave‐Assisted One‐Step Synthesis of 2′,3′‐Cyclic Phosphates of Nucleosides

2023

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Cyclic canonical nucleotides are an important class of compounds playing broad roles in regulating biological processes and are investigated in the context of prebiotic chemistry as activated nucleotides for oligonucleotide formation. Despite their growing importance, synthetic access of 2 ,3 -cyclic nucleotides is constrained, resulting in their cost-prohibitive commercial prices. Here, we describe a microwave-assisted one-pot synthesis starting from commercially available nucleosides employing an easily available cyclophosphorylating reagent, bis(dimethyldiamino)phosphorodiamidate (BDMDAP). The corresponding 2 ,3 -cyclic nucleotides are isolated in good yields (70-91%) by a simple ion-exchange column with no further workup. The nucleosides require no protecting group as the cyclophosphorylation reaction is selective for the 2 ,3 -dihydroxyl groups. The experimental protocol is robust and can be run in parallel to provide access to gram quantities of these 2 ,3 -cyclic nucleotides within a day.

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“…We have so far used a selection of 11 aminoglycoside receptors to distinguish the five canonical RNA motifs: bulges, hairpin loops, stems, symmetrical internal loops, and asymmetrical internal loops. We employ a fluorescence assay with the solvatochromic chemosensor, benzofuranyl uridine (BFU), which was initially incorporated into 16 RNA structures with variable sequences. − The BFU modification is less bulky than other fluorophores, making it possible to modify more sites within compact RNA structures. Using the raw fluorescence data in principal component analysis (PCA), an unbiased statistical method, we were able to classify the five motifs with 100% predictive power and gained insight into the important topological differences that allowed structural differentiation.…”

Section: Introductionmentioning

confidence: 99%

Visualizing RNA Conformational Changes via Pattern Recognition of RNA by Small Molecules

Eubanks¹,

Zhao²,

Thompson³

et al. 2019

J. Am. Chem. Soc.

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Conformational changes in RNA play vital roles in the regulation of many biological systems, yet these changes can be challenging to visualize. Previously, we demonstrated that Pattern Recognition of RNA by Small Molecules (PRRSM) can unbiasedly cluster defined RNA secondary structure motifs utilizing an aminoglycoside receptor library. In this work, we demonstrate the power of this method to visualize changes in folding at the secondary structure level within two distinct riboswitch structures. After labeling at three independent positions on each riboswitch, PRRSM accurately classified all apo and ligand-bound riboswitch structures, including changes in the size of a structural motif, and revealed modification sites that prevented folding and/or led to a mixture of states. These data underscore the utility and robustness of the PRRSM assay for rapid assessment of RNA structural changes and for gaining ready insight into nucleotide positions critical to RNA folding.

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Microwave‐Assisted Phosphitylation of DNA and RNA Nucleosides and Their Analogs

Cited by 4 publications

References 18 publications

Small Molecule-Based Pattern Recognition To Classify RNA Structure

Small Molecule-Based Pattern Recognition To Classify RNA Structure

Microwave‐Assisted One‐Step Synthesis of 2′,3′‐Cyclic Phosphates of Nucleosides

Visualizing RNA Conformational Changes via Pattern Recognition of RNA by Small Molecules

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