Stable isotope labeling for matrix‐assisted laser desorption/ionization mass spectrometry and post‐source decay analysis of ribonucleic acids

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Abstract
Matrix-assisted laser desorption/ionization mass spectrometry is a powerful analytical tool for the structural characterization of oligonucleotides and nucleic acids. Here we report the application of stable isotope labeling for the simplified characterization of ribonucleic acids (RNAs). An 18O label is incorporated at the 3′-phosphate of oligoribonucleotides during the enzymatic processing of intact RNAs. As implemented, a buffer solution containing a 50 : 50 mixture of H2O and 18O-labeled H2O is used during endonuclease digestion. Upon digestion, characteristic doublets representative of the isotopic distribution of oxygen are noted for those products that contain 3′-phosphate groups. This approach is used to distinguish readily endonuclease digestion products from incomplete digestion products and non-specific cleavage products. In addition, RNase digestion products containing the characteristic isotopic doublet can be selected for further characterization by post-source decay (PSD) analysis. PSD products carrying the 3′-phosphate group will appear as a doublet, thereby simplifying fragment ion assignment. Copyright © 2003 John Wiley & Sons, Ltd.