Mesodynamics in the SARS nucleocapsid measured by NMR field cycling

Clarkson, Michael W.; Lei, Ming; Eisenmesser, Elan Z.; Labeikovsky, Wladimir; Redfield, Alfred; Kern, Dorothee

Besides the proof of principle that site-specific relaxation rates can be determined at low field while retaining much of the sensitivity and all of the resolution of a highfield spectrometer, the experiments allowed a precise determination of the internal correlation times of residues with complex motional models (model 5). Given that these residues have very similar internal correlation times and cluster structurally together, we suggest that the relaxation behavior is the result of correlated motions of the hairpin and the 60-64 loop. This model is buttressed by MD simulations. The b-hairpin and the 60-64 loop coincide with several previously-identified sites where SARSN engages in protein-protein and protein-nucleic acid interactions, including a ubiquitination site at K62. That areas of significant flexibility in SARSN coincide with residues important for binding interactions is not surprising, because Fig. 7 The b-hairpin moves collectively as a rigid body and its motion is correlated to motions in adjacent loops. a Representative snapshots from the 15 ns MD trajectory in explicit solvent. The structures are aligned on the globular domain. Trajectory snapshots are blue (5,340 ps) cyan (4,853 ps), orange (10,271 ps), and red (11,033 ps); the original X-ray structure (PDB code: 2OFZ) is in gray. b Time traces of RMSD with respect to the crystal structure. RMSD of the globular portion of the domain when the protein is aligned on the globular domain (as in a) is shown in black, and the RMSD of the hairpin in blue. The RMSD of the hairpin when the protein is aligned on the hairpin is shown in red. c Representative snapshots depicting hydrogen bonds between the side chain of R90 and backbone and side-chain moieties in the 60-64 loop and E129. d Plot showing correlated (red) and anti-correlated (blue) motions of SARSN over the duration of a 15 ns trajectory in a covariance matrix (Ichiye and Karplus 1991) . Motions within the b-hairpin and between the b-hairpin with the 60-64 loop are correlated (marked by circles), in addition to correlated motions between the b-strands in the core of the protein (marked by boxes) disordered regions in proteins are often associated with promiscuous binding activity (Fink 2005 ). However our current data do not provide mechanistic insight into the relation between dynamics and SARS function.