Gradient and sensitivity enhanced multiple-quantum coherence in heteronuclear multidimensional NMR experiments

Recent studies have indicated that the relaxation rate of the ¹H-¹³C multiple-quantum coherence is much slower than that of the ¹H-¹³C single-quantum coherence for non-aromatic methine sites in ¹³ C labeled proteins and in nucleic acids at the slow tumbling limit. Several heteronuclear experiments have been designed to use a multiple-quantum coherence transfer scheme instead of the single-quantum transfer method, thereby increasing the sensitivity and resolution of the spectra. Here, we report a constant time, gradient and sensitivity enhanced HMQC experiment (CT-g/s-HMQC) and demonstrate that it has a significant sensitivity enhancement over constant time HMQC and constant time gradient and sensitivity enhanced HSQC experiments (CT-g/s-HSQC) when applied to a ¹³C and ¹⁵ N labeled calmodulin sample in D₂O. We also apply this approach to 3D NOESY-HMQC and doubly sensitivity enhanced TOCSY-HMQC experiments, and demonstrate that they are more sensitive than their HSQC counterparts.