Non-uniform sampling is shown to provide significant time savings in the acquisition of a suite of three-dimensional NMR experiments utilized for obtaining backbone assignments of H, N, C′, CA, and CB nuclei in proteins: HNCO, HN(CA)CO, HNCA, HN(CO)CA, HNCACB, and HN(CO)CACB. Non-uniform sampling means that data were collected for only a subset of all incremented evolution periods, according to a user-specified sampling schedule. When the suite of six 3D experiments was acquired in a uniform fashion for an 11kDa cytoplasmic domain of a membrane protein at 1.5mM concentration, a total of 146h was consumed. With non-uniform sampling, the same experiments were acquired in 32h and, through subsequent maximum entropy reconstruction, yielded spectra of similar quality to those obtained by conventional Fourier transform of the uniformly acquired data. The experimental time saved with this methodology can significantly accelerate protein structure determination by NMR, particularly when combined with the use of automated assignment software, and enable the study of samples with poor stability at room temperature. Since it is also possible to use the time savings to acquire a greater numbers of scans to increase sensitivity while maintaining high resolution, this methodology will help extend the size limit of proteins accessible to NMR studies, and open the way to studies of samples that suffer from solubility problems.
- Maximum entropy reconstruction
- Non-uniform sampling
- Protein backbone chemical shift assignments
- Reduced time multidimensional NMR spectroscopy
ASJC Scopus subject areas
- Nuclear and High Energy Physics
- Condensed Matter Physics