Integration of spin-state-selective excitation into 2D NMR correlation experiments with heteronuclear ZQ/2Q pi rotations for (1)J(XH)-resolved E.COSY-type measurement of heteronuclear coupling constants in proteins

Spin-State-Selective Excitation ((SE)-E-3), which for example selectively excites amide proton resonances corresponding to exclusively either the alpha or the beta spin slate of the covalently bound N-15 atom is employed for E.COSY-type extraction of heteronuclear J coupling constants. Instead of having one spectrum with two peaks (corresponding to the alpha or beta spin state of N-15), (SE)-E-3 generates two spectra, each with only one peak for each N-15 nucleus. These two spectra are generated from the same data set, so that there is no reduction in sensitivity compared to conventional (1)J(NH)-resolved methods. Another interesting feature in comparison with conventional methods is that (1)J(NH) can be suppressed during the evolution period, meaning that no heteronuclear multiplet structure is visible in the omega(1) frequency dimension. The (SE)-E-3 pulse sequence element is combined with NOESY for measurement of (3)J(N-H)beta and J(N-H)alpha coupling constants in either a hetero- or a homonuclear correlated version. Experimental confirmation is obtained using the protein RAP 17-97 (N-terminal domain of alpha(2)-macroglobulin Receptor Associated Protein).