A PELDOR-based nanometer distance ruler for oligonucleotides

Olav Schiemann, N Piton, YG Mu, G Stock, JW Engels, TF Prisner

Research output: Contribution to journalArticlepeer-review

171 Citations (Scopus)

Abstract

A pulsed electron paramagnetic resonance (EPR) spectroscopic ruler for oligonucleotides was developed using a series of duplex DNAs. The spin-labeling is accomplished during solid-phase synthesis of the oligonucleotides utilizing a palladium-catalyzed cross-coupling reaction between 5-iodo-2'-deoxyuridine and the rigid spin-label 2,2,5,5-tetramethyl-pyrrolin-1-yloxyl-3-acetylene (TPA). 4-Pulse electron double resonance (PELDOR) was then used to measure the intramolecular spin-spin distances via the dipolar coupling, yielding spin-spin distances of 19.2, 23.3, 34.7, 44.8, and 52.5 Angstrom. Employing a full-atom force field with explicit water, molecular dynamic (MD) simulations on the same spin-labeled oligonucleotides in their duplex B-form gave spin-spin distances of 19.6, 21.4, 33.0, 43.3, and 52.5 Angstrom, respectively, in very good agreement with the measured distances. This shows that the oligonucleotides adopt a B-form duplex structure also in frozen aqueous buffer solution. It also demonstrates that the combined use of site-directed spin-labeling, PELDOR experiments, and MD simulations can yield a microscopic picture about the overall structure of oligonucleotides. The technique is also applicable to more complex systems, like ribozymes or DNA/RNA-protein complexes, which are difficult to access by NMR or X-ray crystallography.

Original languageEnglish
Pages (from-to)5722-5729
Number of pages8
JournalJournal of the American Chemical Society
Volume126
Issue number18
DOIs
Publication statusPublished - 12 May 2004

Keywords

  • ELECTRON-SPIN-ECHO
  • PARAMAGNETIC-RESONANCE SPECTROSCOPY
  • HAMMERHEAD RIBOZYME
  • MOLECULAR-DYNAMICS
  • EPR SPECTROSCOPY
  • NUCLEIC-ACIDS
  • PULSE EPR
  • RNA
  • SITE
  • SIMULATION

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