Triple resonance experiments for the simultaneous correlation of H6/H5 and exchangeable protons of pyrimidine nucleotides in 13C,15N-labeled RNA applicable to larger RNA molecules.
Triple-resonance two-dimensional H6/H5(C4N)H and C6/C5(C4N)H experiments are described that provide through-bond H6/H5 or C6/C5 to imino/amino correlations in pyrimidine bases in (13)C,(15)N-labeled RNA. The experiments simultaneously transfer H6/H5 magnetization by an INEPT step to the C6/C5 nuclei and by homonuclear CC- and heteronuclear CN-TOCSY steps via the intervening C4 nucleus to the N3/N4 nuclei and then by a reverse INEPT step to the imino/amino hydrogens. The sensitivity of these experiments is high as demonstrated using a 30-nucleotide pyrimidine rich RNA at a concentration of 0.9 mM at temperatures of 10 degrees C and 25 degrees C. This indicates the general applicability of the experiments and the possibility to obtain correlations for imino resonances in non-canonical regions of the target RNA.
Wöhnert
Jens
J
Institut für Organische Chemie und Chemische Biologie, Center for Biomolecular Magnetic Resonance, Johann Wolfgang Goethe-Universität Frankfurt, Marie-Curie-Str. 11, 60439 Frankfurt/M, Germany. jewoe@nmr.uni-frankfurt.de
Görlach
Matthias
M
Schwalbe
Harald
H
D001483:Base Sequence; D002247:Carbon Isotopes; D007553:Isotope Labeling / Q000379:methods; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D009587:Nitrogen Isotopes; D009690:Nucleic Acid Conformation; D011742:Pyrimidine Nucleotides / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D011742:Pyrimidine Nucleotides; D012313:RNA