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Efficient Synthetic Access to Stable Isotope Labelled Pseudouridine Phosphoramidites for RNA NMR Spectroscopy.
- Source :
-
Chemistry - A European Journal . 6/25/2024, Vol. 30 Issue 36, p1-8. 8p. - Publication Year :
- 2024
-
Abstract
- Here we report the efficient synthetic access to 13C/15N‐labelled pseudouridine phosphoramidites, which were incorporated into a binary H/ACA box guide RNA/product complex comprising 77 nucleotides (nts) in total and into a 75 nt E. coli tRNAGly. The stable isotope (SI) labelled pseudouridines were produced via a highly efficient chemo‐enzymatic synthesis. 13C/15N labelled uracils were produced via chemical synthesis and enzymatically converted to pseudouridine 5′‐monophosphate (ΨMP) by using YeiN, a Ψ‐5′‐monophosphate C‐glycosidase. Removal of the 5′‐phosphate group yielded the desired pseudouridine nucleoside (Ψ), which was transformed into a phosphoramidite building suitable for RNA solid phase synthesis. A Ψ ‐building block carrying both a 13C and a 15N label was incorporated into a product RNA and the complex formation with a 63 nt H/ACA box RNA could be observed via NMR. Furthermore, the SI labelled pseudouridine building block was used to determine imino proton bulk water exchange rates of a 75 nt E. coli tRNAGly CCmnm5U, identifying the TΨC‐loop 5‐methyluridine as a modifier of the exchange rates. The efficient synthetic access to SI‐labelled Ψ building blocks will allow the solution and solid‐state NMR spectroscopic studies of Ψ containing RNAs and will facilitate the mass spectrometric analysis of Ψ‐modified nucleic acids. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 09476539
- Volume :
- 30
- Issue :
- 36
- Database :
- Academic Search Index
- Journal :
- Chemistry - A European Journal
- Publication Type :
- Academic Journal
- Accession number :
- 178095785
- Full Text :
- https://doi.org/10.1002/chem.202401193