Synthesis of (6-13C)Pyrimidine Nucleotides as Spin-Labels for RNA Dynamics

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• 作者：Christoph H. Wunderlich ; Romana Spitzer ; Tobias Santner ; Katja Fauster ; Martin Tollinger ; Christoph Kreutz
• 刊名：The Journal of the American Chemical Society
• 出版年：2012
• 出版时间：May 2, 2012
• 年：2012
• 卷：134
• 期：17
• 页码：7558-7569
• 全文大小：502K
• 年卷期：v.134,no.17(May 2, 2012)
• ISSN：1520-5126

文摘

We present a ¹³C-based isotope labeling protocol for RNA. Using (6-¹³C)pyrimidine phosphoramidite building blocks, site-specific labels can be incorporated into a target RNA via chemical oligonucleotide solid-phase synthesis. This labeling scheme is particularly useful for studying milli- to microsecond dynamics via NMR spectroscopy, as an isolated spin system is a crucial prerequisite to apply Carr鈥揚urcell鈥揗eiboom鈥揋ill (CPMG) relaxation dispersion type experiments. We demonstrate the applicability for the characterization and detection of functional dynamics on various time scales by incorporating the (6-¹³C)uridine and -cytidine labels into biologically relevant RNAs. The refolding kinetics of a bistable terminator antiterminator segment involved in the gene regulation process controlled by the preQb>1b> riboswitch class I was investigated. Using ¹³C CPMG relaxation dispersion NMR spectroscopy, the milli- to microsecond dynamics of the HIV-1 transactivation response element RNA and the Varkud satellite stem loop V motif was addressed.