Expanding the chemical scope of RNA

• Verfasst von: Motorin, Yuri [VerfasserIn]
• Verfasst von: Burhenne, Jürgen [VerfasserIn]
• Verfasst von: Teimer, Roman [VerfasserIn]
• Verfasst von: Koynov, Kaloian [VerfasserIn]
• Verfasst von: Willnow, Sophie [VerfasserIn]
• Verfasst von: Weinhold, Elmar [VerfasserIn]
• Verfasst von: Helm, Mark [VerfasserIn]
• Titel: Expanding the chemical scope of RNA
• Titelzusatz: methyltransferases to site-specific alkynylation of RNA for click labeling
• Verf.angabe: Yuri Motorin, Jürgen Burhenne, Roman Teimer, Kaloian Koynov, Sophie Willnow, Elmar Weinhold and Mark Helm
• Jahr: 2011
• Umfang: 10 S.
• Fussnoten: First published online: 30 October 2010 ; Gesehen am 08.09.2022
• Titel Quelle: Enthalten in: Nucleic acids research
• Ort Quelle: Oxford : Oxford Univ. Press, 1974
• Jahr Quelle: 2011
• Band/Heft Quelle: 39(2011), 5, Seite 1943-1952
• ISSN Quelle: 1362-4962
• DOI: doi:10.1093/nar/gkq825
• Datenträger: Online-Ressource
• Sprache: eng
• K10plus-PPN: 1816299421

Abstract

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.