Stereoselective RNA reaction with chiral 2'-OH acylating agents.
2023; 14 (45): 13235-13243
The reactivity of RNA 2'-OH groups with acylating agents has recently been investigated for high-yield conjugation of RNA strands. To date, only achiral molecules have been studied for this reaction, despite the complex chiral structure of RNA. Here we prepare a set of chiral acylimidazoles and study their stereoselectivity in RNA reactions. Reactions performed with unfolded and folded RNAs reveal that positional selectivity and reactivity vary widely with local RNA macro-chirality. We further document remarkable effects of chirality on reagent reactivity, identifying an asymmetric reagent with 1000-fold greater reactivity than prior achiral reagents. In addition, we identify a chiral compound with higher RNA structural selectivity than any previously reported RNA-mapping species. Further, azide-containing homologs of a chiral dimethylalanine reagent were synthesized and applied to local RNA labeling, displaying 92% yield and 16 : 1 diastereoselectivity. The results establish that reagent stereochemistry and chiral RNA structure are critical elements of small molecule-RNA reactions, and demonstrate new chemical strategies for selective RNA modification and probing.
View details for DOI 10.1039/d3sc03067a
View details for PubMedID 38023505
View details for PubMedCentralID PMC10664579
Sulfonylation of RNA 2'-OH groups.
ACS central science
2023; 9 (3): 531-539
The nucleophilic reactivity of RNA 2'-OH groups in water has proven broadly useful in probing, labeling, and conjugating RNA. To date, reactions selective to ribose 2'-OH have been limited to bond formation with short-lived carbonyl electrophiles. Here we report that many activated small-molecule sulfonyl species can exhibit extended lifetimes in water and retain 2'-OH reactivity. The data establish favorable aqueous solubility for selected reagents and successful RNA-selective reactions at stoichiometric and superstoichiometric yields, particularly for aryl sulfonyltriazole species. We report that the latter are considerably more stable than most prior carbon electrophiles in aqueous environments and tolerate silica chromatography. Furthermore, an azide-substituted sulfonyltriazole reagent is developed to introduce labels into RNA via click chemistry. In addition to high-yield reactions, we find that RNA sulfonylation can also be performed under conditions that give trace yields necessary for structure mapping. Like acylation, the reaction occurs with selectivity for unpaired nucleotides over those in the duplex structure, and a sulfonate adduct causes reverse transcriptase stops, suggesting potential use in RNA structure analysis. Probing of rRNA is demonstrated in human cells, indicating possible cell permeability. The sulfonyl reagent class enables a new level of control, selectivity, versatility, and ease of preparation for RNA applications.
View details for DOI 10.1021/acscentsci.2c01237
View details for PubMedID 36968531
View details for PubMedCentralID PMC10037496
Diverse Reagent Scaffolds Provide Differential Selectivity of 2'-OH Acylation in RNA.
Journal of the American Chemical Society
RNA 2'-OH acylation is widely used both for mapping structure and for conjugating RNA, generally relying on selective reactions with unpaired nucleotides over paired ones. Common reagents for this acylation have been chiefly restricted to two similar aryl scaffolds, leaving open the question of how more broadly varied reagent structure might affect selectivity. Here, we prepared a set of 10 structurally diverse acylimidazole reagents and employed deep sequencing to profile their reactivity and selectivity in an RNA library of systematically varied structure. We show that structure-directed reactivity profiles vary significantly with the reagent scaffold, and we document new acylating agents that have altered selectivity profiles, including reagents that show elevated selectivity within loops, as well as compounds with reduced off-target reactivity in loop closing base pairs. Interestingly, we also show that the simplest reagent (acetylimidazole) is cell permeable and is small enough to map RNA structure in the presence of protein contacts that block other reagents. Finally, we describe reagents that show elevated selectivity within small loops, with applications in site-selective labeling. The results provide new tools for improved conjugation and mapping of RNA.
View details for DOI 10.1021/jacs.2c09040
View details for PubMedID 36542611