Analysis of the ensemble of conformations ofD0
The ensemble of conformations generated by the replica-exchange algorithm at 300K were further analyzed to characterizeD0 . The average Rg , of value 2.19 ± 0.48 nm (cf. Fig. S11), was consistently larger than the value 1.28 nm of the native conformation. The fact that the contact probability between pairs of residues as a function of their distance along the chain is a power law with an exponent ≈-1.8 (cf. Fig. S12) suggests that the chain is, on average, in a coil state.
The average number of contacts is 60.6 ± 15.3 (cf. Fig. S13) and the fraction qN of native contacts is low (≈ 0.018 ± 0.011). To be noted the fraction qN of native contacts in the denatured state is poorly correlated with the commonly employed RMSD (cf. Fig. S14). In fact, the RMSD is a highly non-linear function of the diversity between conformations in that it is very sensitive to conformational changes between similar conformations and quite insensitive to large conformational changes between dissimilar conformations. Since the denatured state is expected to be conformationally very heterogeneous, we compared pairs of conformations using the fraction q of common contacts (and we compared a conformation to the native one using the fractionqN of native contacts).
In Fig. 8(a) the distribution of common contacts q between denatured conformations is plotted. Its average is 0.09 ± 0.08 but it displays a tail up to 0.7. Not surprisingly, the denatured stateD0 thus appears conformationally very heterogeneous. However, its average contact map (cf. Fig. 8(b)) displays well-defined secondary structures that can reach a probability of 0.4 and also tertiary structures populated with probabilities up to 0.15. Some of these structures are native-like and include the hairpin β1-β2, the hairpin β4-β5, the hairpin β5-β6 and the terminal α-helix (cf. Fig. 8(c)). Non-native contacts (cf. Fig. 8(d)) include a set of alternative structures in the region of the hairpin β4-β5, some fluctuating structure around P63 and a small amount of tertiary contacts.