Structural heterogeneity in microcrystalline ubiquitin studied by solid-state NMR

By applying [1-C-13]- and [2-C-13]-glucose labeling schemes to the folded globular protein ubiquitin, a strong reduction of spectral crowding and increase in resolution in solid-state NMR (ssNMR) spectra could be achieved. This allowed spectral resonance assignment in a straightforward manner and the collection of a wealth of long-range distance information. A high precision solid-state NMR structure of microcrystalline ubiquitin was calculated with a backbone rmsd of 1.57 to the X-ray structure and 1.32 angstrom to the solution NMR structure. Interestingly, we can resolve structural heterogeneity as the presence of three slightly different conformations. Structural heterogeneity is most significant for the loop region 1-2 but also for -strands 1, 2, 3, and 5 as well as for the loop connecting 1 and 3. This structural polymorphism observed in the solid-state NMR spectra coincides with regions that showed dynamics in solution NMR experiments on different timescales.