Measuring nanometer distances in nucleic acids using a sequence-independent nitroxide probe.
This protocol describes the procedures for measuring nanometer distances in nucleic acids using a nitroxide probe that can be attached to any nucleotide within a given sequence. Two nitroxides are attached to phosphorothioates that are chemically substituted at specific sites of DNA or RNA. Inter-nitroxide distances are measured using a four-pulse double electron-electron resonance technique, and the measured distances are correlated to the parent structures using a Web-accessible computer program. Four to five days are needed for sample labeling, purification and distance measurement. The procedures described herein provide a method for probing global structures and studying conformational changes of nucleic acids and protein/nucleic acid complexes.
Qin
Peter Z
PZ
Department of Chemistry, University of Southern California, Los Angeles, California 90089-0744, USA. pzq@usc.edu
Haworth
Ian S
IS
Cai
Qi
Q
Kusnetzow
Ana K
AK
Grant
Gian Paola G
GP
Price
Eric A
EA
Sowa
Glenna Z
GZ
Popova
Anna
A
Herreros
Bruno
B
He
Honghang
H
D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D015336:Molecular Probe Techniques; D009589:Nitrogen Oxides / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*; D012984:Software; D013113:Spin Labels
D009589:Nitrogen Oxides; D013113:Spin Labels; D012313:RNA; D004247:DNA; C039900:nitroxyl