Complementing global measures of RNA folding with local reports of backbone solvent accessibility by time resolved hydroxyl radical footprinting.
A variety of analytical techniques are used to probe the mechanisms by which RNA molecules fold to discrete three dimensional structures. Methods such as small angle X-ray scattering (SAXS) report global properties like overall size and shape of the RNA. Other methods such as chemical or enzymatic mapping (footprinting) report properties with resolution as fine as single nucleotide. The hydroxyl radical (*OH) is a footprinting probe which cleaves the oligonucleotide backbone independently of sequence and thus is a valuable reporter of backbone solvent accessibility. Combinations of global and local measures of folding reactions are uniquely able to distinguish specific from nonspecific processes. This article highlights the application of *OH footprinting as a complement to SAXS for kinetics analysis of RNA folding. We illustrate this combination of techniques using a study of the role played by the stiffness of a hinge in determining the rate limiting step of a Mg(2+)-mediated RNA folding reaction.
Schlatterer
Jörg C
JC
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Brenowitz
Michael
M
D001703:Biophysics / Q000379:methods; D004867:Equipment Design; D017665:Hydroxyl Radical; D007700:Kinetics; D015233:Models, Statistical; D009690:Nucleic Acid Conformation; D011108:Polymers / Q000737:chemistry; D012313:RNA / Q000737:chemistry*; D016337:RNA, Catalytic / Q000737:chemistry; D012542:Scattering, Radiation; D053838:Scattering, Small Angle; D012997:Solvents / Q000737:chemistry; D016808:Tetrahymena thermophila / Q000378:metabolism; D013997:Time Factors; D014965:X-Rays
D011108:Polymers; D016337:RNA, Catalytic; D012997:Solvents; D017665:Hydroxyl Radical; D012313:RNA