Identification of 5-hydroxycytidine at position 2501 concludes characterization of modified nucleotides in E. coli 23S rRNA

Jesper Foged Havelund, Anders Michael Bernth Giessing, Trine Møller Hansen, Anette Rasmussen, Lincoln Greyson Scott, Finn Kirpekar

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Complete characterization of a biomolecule's chemical structure is crucial in the full understanding of the relations between their structure and function. The dominating components in ribosomes are ribosomal RNAs (rRNAs), and the entire rRNA-but a single modified nucleoside at position 2501 in 23S rRNA-has previously been characterized in the bacterium Escherichia coli. Despite a first report nearly 20 years ago, the chemical nature of the modification at position 2501 has remained elusive, and attempts to isolate it have so far been unsuccessful. We unambiguously identify this last unknown modification as 5-hydroxycytidine-a novel modification in RNA. Identification of 5-hydroxycytidine was completed by liquid chromatography under nonoxidizing conditions using a graphitized carbon stationary phase in combination with ion trap tandem mass spectrometry and by comparing the fragmentation behavior of the natural nucleoside with that of a chemically synthesized ditto. Furthermore, we show that 5-hydroxycytidine is also present in the equivalent position of 23S rRNA from the bacterium Deinococcus radiodurans. Given the unstable nature of 5-hydroxycytidine, this modification might be found in other RNAs when applying the proper analytical conditions as described here.

Original languageEnglish
JournalJournal of Molecular Biology
Volume411
Issue number3
Pages (from-to)529-536
Number of pages8
ISSN0022-2836
DOIs
Publication statusPublished - 2011

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