Label-free, Direct Localization and Relative Quantitation of the RNA Nucleobase Methylations M6A, M5C, M3U, and M5U by Top-down Mass Spectrometry

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2017 May 27

PMID 28549193

Citations 29

Authors

Heidelinde Glasner

Christian Riml

Ronald Micura

Kathrin Breuker

Affiliations

Soon will be listed here.

Abstract

Nucleobase methylations are ubiquitous posttranscriptional modifications of ribonucleic acids (RNA) that can substantially increase the structural diversity of RNA in a highly dynamic fashion with implications for gene expression and human disease. However, high throughput, deep sequencing does not generally provide information on posttranscriptional modifications (PTMs). A promising alternative approach for the characterization of PTMs, i.e. their identification, localization, and relative quantitation, is top-down mass spectrometry (MS). In this study, we have investigated how specific nucleobase methylations affect RNA ionization in electrospray ionization (ESI), and backbone cleavage in collisionally activated dissociation (CAD) and electron detachment dissociation (EDD). For this purpose, we have developed two new approaches for the characterization of RNA methylations in mixtures of either isomers of RNA or nonisomeric RNA forms. Fragment ions from dissociation experiments were analyzed to identify the modification type, to localize the modification sites, and to reveal the site-specific, relative extent of modification for each site.

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