RAPID MEASUREMENT OF SCALAR 3-BOND H-1(N)-H-1(ALPHA) SPIN COUPLING-CONSTANTS IN N-15-LABELED PROTEINS

Two new 2D NMR experiments, CT-HMQC-HA and CT-HMQC-HN, are proposed fo r the rapid measurement of homonuclear (3)J(HNH alpha) coupling consta nts of uniformly N-15-enriched proteins in solution. The experiments a re based on the comparison of the signal intensities in a pair of cons tant-time [N-15,H-1]-HMQC spectra recorded with and without decoupling of the amide proton - alpha proton coupling. Experimental data record ed with the 78-residue N-terminal domain of the E. coli arginine repre ssor (ArgR-N) and with oxidized E. coli flavodoxin (176 residues) show ed good agreement with 3JHNHa coupling constants obtained by fitting o f the multiplet fine structure of the amide proton resonances or from a 3D HNHA-J experiment, respectively. Quantitative estimates for the e ffects from different relaxation rates of in-phase and antiphase magne tization are given.