Base-type-selective high-resolution 13C edited NOESY for sequential assignment of large RNAs.
Extensive spectral overlap presents a major problem for the NMR study of large RNAs. Here we present NMR techniques for resolution enhancement and spectral simplification of fully 13C labelled RNA. High-resolution 1H-13C correlation spectra are obtained by combining TROSY-type experiments with multiple-band-selective homonuclear 13C decoupling. An additional C-C filter sequence performs base-type-selective spectral editing. Signal loss during the filter is significantly reduced because of TROSY-type spin evolution. These tools can be inserted in any 13C-edited multidimensional NMR experiment. As an example we have chosen the 13C-edited NOESY which is a crucial experiment for sequential resonance assignment of RNA. Application to a 33-nucleotide RNA aptamer and a 76-nucleotide tRNA illustrates the potential of this new methodology.
Brutscher
B
B
Institut de Biologie Structurale-Jean-Pierre Ebel, CNRS-CEA, Grenoble, France.
Boisbouvier
J
J
Kupce
E
E
Tisné
C
C
Dardel
F
F
Marion
D
D
Simorre
J P
JP
D001483:Base Sequence; D002247:Carbon Isotopes; D006859:Hydrogen / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry*; D012343:RNA, Transfer / Q000737:chemistry
D002247:Carbon Isotopes; D012313:RNA; D006859:Hydrogen; D012343:RNA, Transfer