Mesodynamics in the SARS nucleocapsid measured by NMR field cycling

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

The { 1 H}-15 N heteronuclear NOEs for the b-hairpin also display a pronounced field-dependence between 50.7 and 91.2 MHz (Fig. 3) with a confirmation of the previously reported strong NOEs at 50.7 MHz (Huang et al. 2004) . The attenuation of the NOEs at higher fields strongly suggests an internal mesodynamic fluctuation. The 15 N NOE is the ratio R NOE c H /R 1 c N , where R NOE is approximately proportional to J(x H ) since x N ( x H , and R 1 is proportional to J(x N ) (Abragam 1961 ). This ratio changes little for rigid residues over a twofold change in Larmor frequency because the respective spectral density values are essentially constant over this frequency range. In contrast, the observed field-dependence of the NOE for flexible residues indicates the presence of a transition in the spectral density over this frequency range. Moreover, the range of Larmor frequencies sampled defines upper and lower bounds for the internal correlation time s (xs & 1 for the center of the transition) of 150 ps \ s \ 3 ns. Without knowing where the R NOE c H /R 1 c N ratio flattens out, however, the frequency of the motion cannot be determined with great precision. b Expansion of the highfield data. The crossover for the b-hairpin (residues 90-108)-lower R 1 values than the stable and rigid region of the protein at low fields and higher values for fields higher than 50.7 MHz-is apparent Model-free analysis to characterize the dynamics of SARSN While dispersion patterns may imply gross features of the dynamics, a detailed characterization of protein motions requires a more complete survey of relaxation data and interpretation using motional models. For this goal, a complete set of high-field R 1 (Fig. 2b) , NOE (Fig. 3) and R 2 (Fig. 4) data was acquired and used to determine modelfree parameters for all residues of SARSN (Fig. 5) . All residues were fit to a local dynamics model involving either a single order parameter (S 2 ) and internal correlation time (s e ) (model 2, or simple model-free) (Lipari and Szabo 1982) or two order parameters (S 2 f and S 2 s ) and internal correlation times (s s and s f ), where s f is assumed to be *0 (model 5, or extended model-free) (Clore et al. 1990) .