Determination of (3)J(H3(i)', Pi+1) and (3)J(H5(i)'/5(i)'',P-i) coupling constants in C-13-labeled nucleic acids using constant-time HMQC

A novel H-1-C-13 correlated two-dimensional experiment, CT-HMQC-J, for the measurement of three-bond proton-phosphorus coupling constants in C-13-labe led DNA is described, The experiment is based on the intensity difference o f H-1-C-13 cross peaks in the presence and absence of the proton-phosphorus coupling interaction during the constant-time period in HMQC experiment. T he (3)J(H, P) coupling constants can be easily extracted from the intensity ratios of the two experiments. The method has been applied to a uniformly C-13,N-15-labeled d(GGAGGAT) 7-mer DNA sample, The proton-phosphorus coupli ng constants determined from CT-HMQC-J, together with the other three-bond coupling constants, are used to determine beta and epsilon torsion angles. The introduction of beta and epsilon restraints has improved the convergenc e as well as the quality of d(GGAGGAT) structure. (C) 1999 Academic Press.