Synthesis of spin-labeled RNAs for long range distance measurements by PELDOR

Site directed spin labeled RNA duplexes with different interspin distances were synthesized. The radical 2,2,5,5-tetramethyl-pyrrolin-1-yloxyl-3-acetylene (TPA) was introduced during the solid-phase synthesis through a Sonogashira cross-coupling with 5-iodo-uridine. T-m and CD studies showed that the spin label does not to disturb significantly the A-form of these duplexes. 4-Pulse Electron Double Resonance (PELDOR) was then used to measure intramolecular spin-spin distances of 19.3, 33.0 and 40.9 angstrom, which are in very good agreement with the calculated values of 17 6 32.1 and 39.1 angstrom, obtained from Molecular Dynamics (MD) simulations.