C‑Nucleosides Stabilize RNA by Reducing Nucleophilicity at 2'-OH.
Dipanwita Banerjee1, Lu Xiao1, Pavitra S Thacker1
1Department of Chemistry, Stanford University, Stanford, California 94305, United States.
ACS Central Science
|December 31, 2025
Summary
Carbon-substituted nucleotides, like pseudouridine, significantly slow RNA degradation by reducing strand cleavage rates. This discovery offers insights into RNA stability for biological processes and therapeutic applications.
Area of Science:
- Biochemistry
- Molecular Biology
- RNA Therapeutics
Background:
- Carbon-substituted nucleotides are prevalent in biological and therapeutic RNAs, including pseudouridine and N1-methylpseudouridine.
- These modifications are known to enhance RNA stability, but the underlying mechanism remains unclear.
Purpose of the Study:
- To investigate the mechanism by which carbon-substituted nucleotides reduce RNA degradation.
- To elucidate the impact of carbon substitution on RNA cleavage kinetics and nucleophilicity.
Main Methods:
- Kinetic analysis of spontaneous and enzymatic cleavage at single bonds in synthetically modified RNAs.
- Studies on nucleophilic acylation reactions involving modified RNAs and alcohols with varying pKa.
Main Results:
- Carbon substitution significantly decreases RNA strand cleavage rates via both spontaneous and enzymatic pathways.
- Reduced inductive effects from carbon substitution lead to higher pKa and lower nucleophilicity in RNA.
Conclusions:
- Carbon substitution in nucleotides enhances RNA stability by reducing cleavage rates.
- The findings provide a mechanistic understanding of RNA modifications relevant to native transcriptomes and RNA-based therapies.
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