IMPROVED SENSITIVITY AND RESOLUTION IN H-1-C-13 NMR EXPERIMENTS OF RNA

NMR studies of the structure and dynamics in RNA are greatly facilitat ed by the use of C-13-labeled molecules and H-1-C-13 correlation exper iments. We demonstrate here that spin-state selective C-13 frequency e diting yields greatly improved sensitivity and resolution in H-1-C-13 correlation spectra of RNA. A sensitivity enhanced version of the prev iously introduced TROSY sequence (Pervushin et al., Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 12366-12371) is proposed, which offers a root 2 signal enhancement. When compared to a sensitivity enhanced HSQC seque nce an up to 3-fold increase in both the sensitivity and resolution is observed for the base carbons in a 15% C-13-labeled 33-mer RNA at a m agnetic field strength of 14.1 T. This increase results from the relax ation interference between the C-13 chemical shielding anisotropy (CSA ) and the H-1-C-13 dipolar interaction, as well as the simultaneous de tection of H-1 and C-13 steady-state polarizations. Simulations indica te that the enhancement effect is maximal at currently available magne tic field strengths (less than or equal to 18.8 T). It is further show n that the recording of a pair of complementary TROSY experiments, den oted H-(alpha)- and H-(beta)-TROSY allows the accurate measurement of H-1-C-13 one-bond coupling constants from the cross-peak positions in the two spectra. An extension of the TROSY sequence is presented for r ecording high resolution 3D C-13-edited NOESY spectra.