NMR spectroscopy of RNA duplexes containing pseudouridine in supercooled water.
We have performed NMR experiments in supercooled water in order to decrease the temperature-dependent exchange of protons in RNA duplexes. NMR spectra of aqueous samples of RNA in bundles of narrow capillaries that were acquired at temperatures as low as -18 degrees C reveal resonances of exchangeable protons not seen at higher temperatures. In particular, we detected the imino protons of terminal base pairs and the imino proton of a non-base-paired pseudouridine in a duplex representing the eukaryotic pre-mRNA branch site helix. Analysis of the temperature dependence of chemical shift changes (thermal coefficients) for imino protons corroborated hydrogen bonding patterns observed in the NMR-derived structural model of the branch site helix. The ability to observe non-base-paired imino protons of RNA is of significant value in structure determination of RNA motifs containing loop and bulge regions.
Schroeder
Kersten T
KT
Department of Chemistry and Biochemistry, Dittmer Laboratory of Chemistry, Florida State University, Tallahassee, FL 32306-4390, USA.
Skalicky
Jack J
JJ
Greenbaum
Nancy L
NL
D001483:Base Sequence; D003080:Cold Temperature; D006860:Hydrogen Bonding; D008956:Models, Chemical; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D011522:Protons; D011560:Pseudouridine / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D014867:Water / Q000737:chemistry*
D011522:Protons; D014867:Water; D011560:Pseudouridine; D012313:RNA