Quick Access to Nucleobase-Modified Phosphoramidites for the Synthesis of Oligoribonucleotides Containing Post-Transcriptional Modifications and Epitranscriptomic Marks

Overview

Journal J Org Chem

Publisher American Chemical Society

Specialty Chemistry

Date 2022 Jul 20

PMID 35857285

Authors

Kamil Ziemkiewicz

Marcin Warminski

Radoslaw Wojcik

Joanna Kowalska

Jacek Jemielity

Affiliations

Soon will be listed here.

Abstract

Herein, we report a straightforward one-step procedure for modifying -nucleophilic groups in the nucleobases of commercially available nucleoside phosphoramidites. This method involves the deprotonation of amide groups under phase-transfer conditions and subsequent reaction with electrophilic molecules such as alkyl halides or organic isocyanates. Using this approach, we obtained 10 different classes of modified nucleoside phosphoramidites suitable for the synthesis of oligonucleotides, including several noncanonical nucleotides found in natural RNA or DNA (e.g., mA, iA, mA, gA, mC, mC, mU, mG, and mG). Such modification of nucleobases is a common mechanism for post-transcriptional regulation of RNA stability and translational activity in various organisms. To better understand this process, relevant cellular recognition partners (e.g., proteins) must be identified and characterized. However, this step has been impeded by limited access to molecular tools containing such modified nucleotides.

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References

Lewdorowicz M, Yoffe Y, Zuberek J, Jemielity J, Stepinski J, Kierzek R . Chemical synthesis and binding activity of the trypanosomatid cap-4 structure. RNA. 2004; 10(9):1469-78. PMC: 1370633. DOI: 10.1261/rna.7510504. View

Nainyte M, Muller F, Ganazzoli G, Chan C, Crisp A, Globisch D . Amino Acid Modified RNA Bases as Building Blocks of an Early Earth RNA-Peptide World. Chemistry. 2020; 26(65):14856-14860. PMC: 7756884. DOI: 10.1002/chem.202002929. View

Nainyte M, Amatov T, Carell T . Synthesis of an acpU phosphoramidite and incorporation of the hypermodified base into RNA. Chem Commun (Camb). 2019; 55(81):12216-12218. DOI: 10.1039/c9cc06314e. View

Murphy 4th F, Ramakrishnan V, Malkiewicz A, Agris P . The role of modifications in codon discrimination by tRNA(Lys)UUU. Nat Struct Mol Biol. 2004; 11(12):1186-91. DOI: 10.1038/nsmb861. View

Harcourt E, Kietrys A, Kool E . Chemical and structural effects of base modifications in messenger RNA. Nature. 2017; 541(7637):339-346. PMC: 5498787. DOI: 10.1038/nature21351. View

McCown P, Ruszkowska A, Kunkler C, Breger K, Hulewicz J, Wang M . Naturally occurring modified ribonucleosides. Wiley Interdiscip Rev RNA. 2020; 11(5):e1595. PMC: 7694415. DOI: 10.1002/wrna.1595. View

Sood A, Viner C, Hoffman M . DNAmod: the DNA modification database. J Cheminform. 2019; 11(1):30. PMC: 6478773. DOI: 10.1186/s13321-019-0349-4. View

Griffey R, Monia B, Cummins L, Freier S, Greig M, Guinosso C . 2'-O-aminopropyl ribonucleotides: a zwitterionic modification that enhances the exonuclease resistance and biological activity of antisense oligonucleotides. J Med Chem. 1996; 39(26):5100-9. DOI: 10.1021/jm950937o. View

Xu C, Liu K, Ahmed H, Loppnau P, Schapira M, Min J . Structural Basis for the Discriminative Recognition of N6-Methyladenosine RNA by the Human YT521-B Homology Domain Family of Proteins. J Biol Chem. 2015; 290(41):24902-13. PMC: 4598999. DOI: 10.1074/jbc.M115.680389. View

10.

Sednev M, Mykhailiuk V, Choudhury P, Halang J, Sloan K, Bohnsack M . N -Methyladenosine-Sensitive RNA-Cleaving Deoxyribozymes. Angew Chem Int Ed Engl. 2018; 57(46):15117-15121. DOI: 10.1002/anie.201808745. View

11.

Hagelskamp F, Borland K, Ramos J, Hendrick A, Fu D, Kellner S . Broadly applicable oligonucleotide mass spectrometry for the analysis of RNA writers and erasers in vitro. Nucleic Acids Res. 2020; 48(7):e41. PMC: 7144906. DOI: 10.1093/nar/gkaa091. View

12.

Liaqat A, Stiller C, Michel M, Sednev M, Hobartner C . N -Isopentenyladenosine in RNA Determines the Cleavage Site of Endonuclease Deoxyribozymes. Angew Chem Int Ed Engl. 2020; 59(42):18627-18631. PMC: 7589339. DOI: 10.1002/anie.202006218. View

13.

Verdolino V, Cammi R, Munk B, Schlegel H . Calculation of pKa values of nucleobases and the guanine oxidation products guanidinohydantoin and spiroiminodihydantoin using density functional theory and a polarizable continuum model. J Phys Chem B. 2008; 112(51):16860-73. DOI: 10.1021/jp8068877. View

14.

McKenzie L, El-Khoury R, Thorpe J, Damha M, Hollenstein M . Recent progress in non-native nucleic acid modifications. Chem Soc Rev. 2021; 50(8):5126-5164. DOI: 10.1039/d0cs01430c. View

15.

Scheitl C, Ghaem Maghami M, Lenz A, Hobartner C . Site-specific RNA methylation by a methyltransferase ribozyme. Nature. 2020; 587(7835):663-667. PMC: 7116789. DOI: 10.1038/s41586-020-2854-z. View

16.

Theler D, Dominguez C, Blatter M, Boudet J, Allain F . Solution structure of the YTH domain in complex with N6-methyladenosine RNA: a reader of methylated RNA. Nucleic Acids Res. 2014; 42(22):13911-9. PMC: 4267619. DOI: 10.1093/nar/gku1116. View

17.

Stuart J, Gdaniec Z, Guenther R, Marszalek M, Sochacka E, Malkiewicz A . Functional anticodon architecture of human tRNALys3 includes disruption of intraloop hydrogen bonding by the naturally occurring amino acid modification, t6A. Biochemistry. 2000; 39(44):13396-404. DOI: 10.1021/bi0013039. View

18.

Shi H, Wei J, He C . Where, When, and How: Context-Dependent Functions of RNA Methylation Writers, Readers, and Erasers. Mol Cell. 2019; 74(4):640-650. PMC: 6527355. DOI: 10.1016/j.molcel.2019.04.025. View

19.

Zhang X, Wei L, Wang Y, Xiao Y, Liu J, Zhang W . Structural insights into FTO's catalytic mechanism for the demethylation of multiple RNA substrates. Proc Natl Acad Sci U S A. 2019; 116(8):2919-2924. PMC: 6386707. DOI: 10.1073/pnas.1820574116. View

20.

Moreno S, Flemmich L, Micura R . Synthesis of -acetylated 3-methylcytidine phosphoramidites for RNA solid-phase synthesis. Monatsh Chem. 2022; 153(3):285-291. PMC: 8948120. DOI: 10.1007/s00706-022-02896-x. View