Residual dipolar (1)H-(1)H couplings of methyl groups in weakly aligned proteins.

Residual dipolar couplings measured for weakly aligned proteins provide important restraints for molecular structure determinations by NMR1 spectroscopy which cannot be obtained otherwise.2 Residual dipolar couplings are usually measured by comparing multiplet splittings measured in anisotropic phase with those measured in isotropic phase.2,3 In the absence of scalar couplings, a residual dipolar coupling between two spins in the weak-coupling limit is directly manifested in a doublet splitting, but the sign of the coupling is more difficult to determine.4,5 Here we show that the sign and magnitude of residual dipolar couplings between the protons of a methyl group are readily measured in a single experiment. These resulting splittings are larger than those due to intra-methyl residual dipolar couplings between 13C and 1H spins, and they depend on the molecular alignment tensor in a way completely analogous to residual dipolar couplings in twospin systems.6 They are thus straightforward to use as structural parameters. Dipolar couplings lead to line splittings even for isolated methyl groups.7 The dipolar contribution to the 1H NMR spectrum of an isolated methyl group is determined by the secular part of the dipolar Hamiltonian which can be decomposed into products of spatial and spin terms:8