, and LEU73-C δ,2 . It appears that the CCR rates of these methyls are systematically overestimated in the MD simulations. This is especially the case for AMBER force field, where all 7 rates are overestimated, whereas CHARMM36 only overestimates 4 rates, LEU8-C δ,2 , VAL70-C γ,2 , LEU71-C δ,1 , and LEU73-C δ,2 . Figure shows that all these methyls display marked internal dynamics with a slow phase in the TCFs, which is due to interconversion between rotamer states of the side chain dihedrals and is also manifested in low order parameters (S 2 axis < 0.4, Table ). The systematic overestimation of the CCR rates of these flexible methyl groups in the MD simulations could thus, at least partly, be attributed to sampling limitations, because even the accumulated simulation time of 1 µs, which exceeds τ c by a factor of 240, might not be sufficient to obtain completely equilibrated Boltzmann distributions of all rotamer states, trans, gauche(+), and gauche(-), of each side chain χ dihedral. As shown below for VAL70, such incomplete rotamer averaging decreases the agreement between MD and NMR especially if the simulations fail to sample rotamer states that display jump dynamics on time scales faster than τ c (and thus contribute to spin relaxation), or if the rotamer transitions are sampled in MD but are too slow, such that the contribution of these motions to relaxation are truncated by overall tumbling. The latter may be especially pronounced for small proteins with fast tumbling times, such as UBQ with τ c = 4.16 ns. This might also be linked to the fact that all MD simulation trajectories were started from the same (single) conformation, as is present in the X-ray crystal structure. Thus, alternative rotamer states, which could well be populated in the NMR ensemble, are not present at the beginning of the simulations but need to be populated through barrier crossings, which could create sampling bottlenecks. Using different conformations as starting structures can be advantageous in such cases, but it should be kept in mind that sampling of the very large configuration space of proteins remains a major challenge. Enhanced sampling methods are essential to more completely explore configuration space. However, while such methods can provide more reliable estimates of the amplitudes of the motions, and thus also of related thermodynamic properties such as conformational entropy, 30,68 the extraction of motional time scales from such biased simulations is challenging. To more deeply understand the conformational dynamics underlying the side chain CCR rates, we computed the populations of the χ 1 dihedral angle rotamer states of the VAL, ILE, and THR residues in ubiquitin from the MD simulations and compared them to the respective populations derived from measurements of 3 J C -Cγ and 3 J N -Cγ couplings and dipolar couplings by Chou and coworkers. Table shows that in general, the agreement is very good for both force fields used, in line with previous findings. 72 Furthermore, these data provide a means to grasp the systematic overestimation of the slow CCR rates of mobile methyl groups (see above). VAL70 serves to illustrate this point. The experimental CCR rate and S 2 axis order parameter of VAL70-C γ,2 are 1.08 s -1 and 0.39, respectively, indicating pronounced side chain mobility. This is in line with the rotamer distribution derived from the experimental couplings, which indicates that both the trans and the gauche(-) states are significantly populated (36% and 59%, respectively), with a minor contribution also of gauche(+). The fact that the gauche(-) minimum is the major conformer is noteworthy because in the X-ray crystal structure, the χ 1 dihedral of VAL70 adopts the trans conformation (Table , last column). Interestingly, the simulations with the AMBER force field provide the same qualitative picture as the NMR experiments, although the population of the gauche(-) conformer is somewhat overestimated (77%) and that of the trans conformer is slightly underestimated (21%). In line with these small deviations, the CCR rate and S 2 axis order parameter of VAL70-C γ,2 from the AMBER simulations are 1.56 s -1 and 0.43, respectively, which are only slightly too high compared to the experimental values. The χ 1 dihedral time trace shown in Figure supports the notion of a mobile VAL70 side chain that undergoes jump-like χ 1 transitions on the nanosecond time scale, that is, on time scales faster than τ c .