Studying RNA using site-directed spin-labeling and continuous-wave electron paramagnetic resonance spectroscopy.
In site-directed spin-labeling (SDSL), a stable nitroxide radical is attached to a specific location within a macromolecule and electron paramagnetic resonance (EPR) spectroscopy is used to interrogate the local environment surrounding the nitroxide. The SDSL strategy enables probing site-specific structural and dynamic features of RNA in solution without being limited by the size of the molecule, thus serving as a unique tool in biophysical studies of RNA. This chapter describes the use of continuous-wave (cw)-EPR to study dynamic features of RNAs as well as to monitor interactions between them. Various approaches for attaching nitroxide spin labels to nucleic acids are described, followed by detailed descriptions of cw-EPR spectral acquisition and processing procedures. Specific examples are subsequently used to illustrate analysis of EPR spectra, showing how information regarding the parent RNA can be extracted.
Zhang
Xiaojun
X
Department of Chemistry, University of Southern California, Los Angeles, California, USA.
Cekan
Pavol
P
Sigurdsson
Snorri Th
ST
Qin
Peter Z
PZ
D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D012313:RNA / Q000737:chemistry*; D013113:Spin Labels
D013113:Spin Labels; D012313:RNA