Amine‐to‐Azide Conversion on Native RNA via Metal‐Free Diazotransfer Opens New Avenues for RNA Manipulations

Abstract

A major challenge in the field of RNA chemistry is the identification of selective and quantitative conversion reactions on RNA that can be used for tagging and any other RNA tool development. Here, we introduce metal-free diazotransfer on native RNA containing an aliphatic primary amino group using the diazotizing reagent fluorosulfuryl azide (FSO 2 N 3). The reaction provides the corresponding azidemodified RNA in nearly quantitatively yields without affecting the nucleobase amino groups. The obtained azido-RNA can then be further processed utilizing well-established bioorthogonal reactions, such as azide-alkyne cycloadditions (Click) or Staudinger ligations. We exemplify the robustness of this approach for the synthesis of peptidyl-tRNA mimics and for the pull-down of 3-(3-amino-3-carboxypropyl)uridine (acp 3 U)-and lysidine (k 2 C)-containing tRNAs of an Escherichia coli tRNA pool isolated from cellular extracts. Our approach therefore adds a new dimension to the targeted chemical manipulation of diverse RNA species.

Azide‐Modified Nucleosides as Versatile Tools for Bioorthogonal Labeling and Functionalization

“…An interesting approach has recently been described by Micura and co-workers, where aliphatic amino groups in RNA sequences were chemically converted into azido moieties (Figure 8A). [94] For this purpose, the new diazotizing reagent fluorosulfuryl azide (FSO 2 N 3 ) was used. [95] Diazotizing reagents for use with ONs have been described before, [96,97] however in these cases the diazotizing reagents were used in a high excess (20 eq.…”

“…The azido functionalities were then used to introduce affinity tags, which allowed for subsequent pull-down of the target RNAs (Figure 8 B). [94] Figure 7. Azido labeling of ONs.…”

“…This allows for a subsequent pulldown of the modified tRNA with streptavidin magnetic beads (blue). [94] Figure 9. DNA with triazole linkages.…”

Azido Functionalized Nucleosides Linked to Controlled Pore Glass as Suitable Starting Materials for Oligonucleotide Synthesis by the Phosphoramidite Approach

“…[13] Owing to this reactivity, azido modifications are often inserted into oligonucleotides enzymatically, [14,15] or chemically after solid phase ON synthesis has been concluded. [16][17][18] H-phosphonate or phosphotriester chemistry was also used instead of the phosphoramidite approach for the synthesis of azido-modified oligonucleotides, [19][20][21] or synthesis of 3'-terminal azido modified ODNs was conducted in reverse orientation from 5' to 3'-end. [10] Yet, there have been reports on the chemical synthesis of azido modified ONs by the standard phosphoramidite strategy.…”

Trendbericht Organische Chemie 2022

“…Mitarbeiter:innen azidomodifizierte RNAs über einen metallfreien Diazotransfer her. 145) Mit primären Aminen modifizierte RNAs lassen sich damit leicht mit Aziden funktionalisieren, sodass sie sich für bioorthogonale Markierungsreaktionen wie die Azid-Alkin-Cycloadditionen oder die Staudinger-Ligation eignen.…”