Chemical-shift anisotropy measurements of amide and carbonyl resonances in a microcrystalline protein with slow magic-angle spinning NMR spectroscopy

Chemical shifts are fundamental to interpretation of NMR spectra and provide constraints for macromolecular structure determination, refinement, and validation, as well as details of active-site chemistry in enzymes. Insights into the origins of the chemical shift can be leveraged to improve chemical analysis, including conformation, bonding, and dynamics. To exploit this information fully, it is desirable to measure not only isotropic chemical shifts but also the full chemical-shift anisotropy (CSA) tensor. Until recently, most efforts to measure backbone amide and carbonyl CSAs have relied upon cross-correlated relaxation and residual anisotropic shifts in solution NMR. In solid-state NMR, analysis of sideband patterns has typically been reserved for site-specifically labeled samples or small peptides. Here we demonstrate that the Herzfeld-Berger method (Herzfeld, J.; Berger, A. E. J. Chem. Phys. 1980, 73, 6021-6030) can be applied to highly C-13,N-15-enriched solid proteins, using 2D heteronuclear correlation in combination with high magnetic fields (750 MHz H-1 frequency) and pattern labeling of C-13 sites. The experiments report on 42 pairs of amide and carbonyl tensors in the microcrystalline protein GB1.