Expanding the chemical scope of RNA: methyltransferases to site-specific alkynylation of RNA for click labeling
This work identifies the combination of enzymatic transfer and click labeling as an efficient method for the site-specific tagging of RNA molecules for biophysical studies. A double-activated analog of the ubiquitous co-substrate S -adenosyl- l -methionine was employed to enzymatically transfer a fi...
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| Main Authors: | , , , , , , |
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| Format: | Article (Journal) |
| Language: | English |
| Published: |
2011
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| In: |
Nucleic acids research
Year: 2011, Volume: 39, Issue: 5, Pages: 1943-1952 |
| ISSN: | 1362-4962 |
| DOI: | 10.1093/nar/gkq825 |
| Online Access: | Verlag, lizenzpflichtig, Volltext: https://doi.org/10.1093/nar/gkq825
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| Author Notes: | Yuri Motorin, Jürgen Burhenne, Roman Teimer, Kaloian Koynov, Sophie Willnow, Elmar Weinhold and Mark Helm |
| Summary: | This work identifies the combination of enzymatic transfer and click labeling as an efficient method for the site-specific tagging of RNA molecules for biophysical studies. A double-activated analog of the ubiquitous co-substrate S -adenosyl- l -methionine was employed to enzymatically transfer a five carbon chain containing a terminal alkynyl moiety onto RNA. The tRNA:methyltransferase Trm1 transferred the extended alkynyl moiety to its natural target, the N2 of guanosine 26 in tRNA Phe . LC/MS and LC/MS/MS techniques were used to detect and characterize the modified nucleoside as well as its cycloaddition product with a fluorescent azide. The latter resulted from a labeling reaction via Cu(I)-catalyzed azide-alkyne 1,3-cycloaddition click chemistry, producing site-specifically labeled RNA whose suitability for single molecule fluorescence experiments was verified in fluorescence correlation spectroscopy experiments. |
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| Item Description: | First published online: 30 October 2010 Gesehen am 08.09.2022 |
| Physical Description: | Online Resource |
| ISSN: | 1362-4962 |
| DOI: | 10.1093/nar/gkq825 |