Kerstin Riedel

MAS solid state NMR of RNAs with multiple receivers.

In the study of biomolecular systems, magic angle spinning (MAS) solid state NMR is emerging as a powerful complementary tool to X-ray crystallography and solution state NMR. Making use of the distance and torsion angle constraints extracted in the solid state the structural characteristics of many biologically interesting systems have been elucidated via MAS solid state NMR recently (Castellani et al. 2002; Jaroniec et al. 2002; Rienstra et al. 2002; Luca et al. 2003; Tycko 2003; Krabben et al. 2004; Zech et al. 2005; Iwata et al. 2006; Egawa et al. 2007; Goldbourt et al. 2007). Homoand heteronuclear distances involving C and N nuclei with well resolved isotropic chemical shifts are commonly used in MAS solid state NMR based structural studies. Although H resonances are generally broad due to strong homonuclear dipolar couplings, the possibilities for extracting short range H–H distance estimates from fully protonated (C, N) labelled peptide/protein samples have been demonstrated recently (de Boer et al. 2002; Lange et al. 2002, 2003, 2005; Tycko and Ishii 2003; Reif et al. 2003). This approach exploits the improved spectral resolution seen in N and C spectra and involves H–H dipolar coupling mediated chemical shift correlation of the low c nuclei. Cross-peak intensities seen in such data, commonly referred to as CHHC, CHHN, NHHN and NHHC spectra, are related to the spatial proximity of the protons that are directly attached to the corresponding nuclei observed in the two dimensions. Our recent studies (Riedel et al. 2005a, 2006) indicate that NHHN, CHHC and NHHC type experiments also hold considerable potential in the structural studies of RNAs that play a critical role in many biological processes and exhibit a variety of secondary and tertiary structural features. Duplex regions arising from consecutive formation of hydrogen bonded base pairs are commonly found in RNA. Hence, the identification of the hydrogen bonded base-pairs and the characterisation of the underlying hydrogen bonding patterns is of critical importance in the study of RNA. As different canonical and non-canonical base-pairing schemes encountered in nucleic acids are characterised by topologically different networks of strong proton–proton dipolar couplings, it has been demonstrated that the characterisation of the hydrogen bonding networks in RNAs can be effectively carried out via NHHN and NHHC type of experiments (Riedel et al. 2005a). In addition, it has also been shown that H–H dipolar coupling mediated C–C chemical shift correlation experiments can facilitate the characterisation of the glycosidic torsion angle v, the sugar pucker and the helical regions of RNAs (Riedel et al. 2006). Hence, data from NHHN, CHHC and NHHC type experiments are critically important in RNA structural studies. Currently, the different proton–proton dipolar coupling mediated N/C chemical shift correlation experiments are carried out individually and, hence, considerable amount of spectrometer time is required to generate data with good signalto-noise ratio. In this communication an efficient approach for the simultaneous collection of these different MAS solid state NMR data sets is presented. The efficacy of the approach is demonstrated using an RNA composed of 97 CUG repeats, (CUG)97, a system that is under investigation Electronic supplementary material The online version of this article (doi:10.1007/s10858-008-9247-1) contains supplementary material, which is available to authorized users.