Journal
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
Volume 59, Issue 20, Pages 7891-7896Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/anie.201916447
Keywords
EPR spectroscopy; PELDOR; RNA; spin labeling; unnatural base pairs
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Funding
- Deutsche Forschungsgemeinschaft [KA 3699/6-1]
- Boehringer Ingelheim Stiftung
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We present herein a novel nitroxide spin label-containing RNA triphosphate TPT3(NO) and its application for site-specific spin-labeling of RNA through in vitro transcription using an expanded genetic alphabet. Our strategy allows the facile preparation of spin-labeled RNAs with sizes ranging from short RNA oligonucleotides to large, complex RNA molecules with over 370 nucleotides by standard in vitro transcription. As a proof of concept, inter-spin distance distributions are measured by pulsed electron paramagnetic resonance (EPR) spectroscopy in short self-complementary RNA sequences and in a well-studied 185 nucleotide non-coding RNA, the B. subtilis glmS ribozyme. The approach is then applied to probe for the first time the folding of the 377 nucleotide A-region of the long non-coding RNA Xist, by PELDOR.
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