Long-range distance determination in fully deuterated RNA with pulsed EPR spectroscopy

Burkhard Endeward, Yanping Hu, Guangcan Bai, Guoquan Liu, Thomas F. Prisner, Xianyang Fang

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

Pulsed electron-electron double resonance (PELDOR or DEER) spectroscopy is powerful in structure and dynamics study of biological macromolecules by providing distance distribution information ranging from 1.8 to 6 nm, providing that the biomolecules are site-specifically labeled with paramagnetic tags. However, long distances up to 16 nm have been measured on perdeuterated and spin-labeled proteins in deuterated solvent by PELDOR. Here we demonstrate long-range distance measurement on a large RNA, the 97-nucleotide 3′SL RNA element of the Dengue virus 2 genome, by combining a posttranscriptional site-directed spin labeling method using an unnatural basepair system with RNA perdeuteration by enzymatic synthesis using deuterated nucleotides. The perdeuteration removes the coupling of the electron spins of the nitroxide spin labels from the proton nuclear spin system of the RNA and does extend the observation time windows of PELDOR up to 50 μs. This enables one to determine long distances up to 14 nm for large RNAs and their conformational flexibility.

Original languageEnglish (US)
Pages (from-to)37-43
Number of pages7
JournalBiophysical journal
Volume121
Issue number1
DOIs
StatePublished - Jan 4 2022
Externally publishedYes

ASJC Scopus subject areas

  • Biophysics

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