Measurement of methyl 13C-1H cross-correlation in uniformly 13C-, 15N-, labeled proteins

An understanding of side chain motions in protein is of great interest since side chains often play an important role in protein folding and intermolecular interactions. A novel method for measuring the dynamics of methyl groups in uniformly C-13-, N-15-labeled proteins has been developed by our group. The method relies on the difference in peak intensities of C-13 quartet components of methyl groups, in a spectrum recording the free evolution of C-13 under proton coupling in a constant-time period. Cross-correlated relaxation rates between C-13-H-1 dipoles can be easily measured from the intensities of the multiplet components. The degree of the methyl restrictions (S-2) can be estimated from the cross-correlated relaxation rate. The method is demonstrated on a sample of human fatty acid binding protein in the absence of fatty acid. We obtained relaxation data for 33 out of 46 residues having methyl groups in apo-IFABP. It has been found that the magnitude of the CSA tensor of spin C-13 in a methyl group could be estimated from the intensities of the C-13 multiplet components.