Chemical Amination/Imination of Carbonothiolated Nucleosides During RNA Hydrolysis

Angew Chem Int Ed Engl. 2021 Feb 19;60(8):3961-3966. doi: 10.1002/anie.202010793. Epub 2020 Dec 10.

Abstract

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has become the gold-standard technique to study RNA and its various modifications. While most research on RNA nucleosides has been focused on their biological roles, discovery of new modifications remains of interest. With state-of-the-art technology, the presence of artifacts can confound the identification of new modifications. Here, we report the characterization of a non-natural mcm5 isoC ribonucleoside in S. cerevisiae total tRNA hydrolysate by higher-energy collisional dissociation (HCD)-based fingerprints and isotope labeling of RNA. Its discovery revealed a class of amino/imino ribonucleoside artifacts that are generated during RNA hydrolysis under ammonium-buffered mild basic conditions. We then identified digestion conditions that can reduce or eliminate their formation. These finding and method enhancements will improve the accurate detection of new RNA modifications.

Keywords: LC-MS/MS; amination; digestion artifact; imination; nucleoside analysis.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amination
  • Chromatography, High Pressure Liquid
  • Hydrolysis
  • Isotope Labeling
  • Nucleosides / chemistry*
  • RNA / analysis*
  • RNA / metabolism
  • RNA, Transfer / chemistry
  • RNA, Transfer / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Sulfhydryl Compounds / chemistry*
  • Tandem Mass Spectrometry

Substances

  • Nucleosides
  • Sulfhydryl Compounds
  • RNA
  • RNA, Transfer