More line narrowing in TROSY by decoupling of long-range couplings:: shift correlation and 1JNc′ coupling constant measurements

Since the introduction of RDCs in high-resolution NMR studies of macromolecules, there is a growing interest in the development of accurate, and sensitive methods for determining coupling constants. Most methods for extracting these couplings are based on the measurement of the splitting between multiplet components in J-coupled spectra. However, these methods are often unreliable since undesired multiple-bond couplings can considerably broaden the multiplet components and consequently make accurate determination of their position difficult. To demonstrate one approach to this problem, G-BIRD(r) decoupled TROSY sequences are proposed for the measurement of (1)J(NH) and (1)J(NC') Coupling constants. Resolved or unresolved splittings due to remote protons are removed by a G-BIRD(r) module employed during t(j) and as a result, spectra with narrow, well-resolved peaks are obtained from which heteronuclear one-bond couplings can be accurately measured. Moreover, introduction of a spin-state-selective alpha/beta-filter in the TROSY sequence allows the separation of the (1)j(NC') doublet components into two subspectra which contain the same number of peaks as the regular TROSY spectrum. The (1)J(NC') couplings are obtained from the displacement between the corresponding peaks in the subspectra. (C) 2004 Elsevier Inc. All rights reserved.