Robustness of atomistic Go models in predicting native-like folding intermediates.

Go models are exceedingly popular tools in computer simulations of protein folding. These models are native-centric, i.e., they are directly constructed from the protein's native structure. Therefore, it is important to understand up to which extent the atomistic details of the native structure dictate the folding behavior exhibited by Go models. Here we address this challenge by performing exhaustive discrete molecular dynamics simulations of a Go potential combined with a full atomistic protein representation. In particular, we investigate the robustness of this particular type of Go models in predicting the existence of intermediate states in protein folding. We focus on the N47G mutational form of the Spc-SH3 folding domain (x-ray structure) and compare its folding pathway with that of alternative native structures produced in silico. Our methodological strategy comprises equilibrium folding simulations, structural clustering, and principal component analysis.

Identification of a conserved aggregation-prone intermediate state in the folding pathways of Spc-SH3 amyloidogenic variants.

We compared the folding pathways of selected mutational variants of the α-spectrin SH3 domain (Spc-SH3) by using a continuum model that combines a full atomistic protein representation with the Go potential. Experimental data show that the N47G mutant shows very little tendency to aggregate while the N47A and triple mutant D48G(2Y) are both amyloidogenic, with the latter being clearly more aggregation prone. We identified a strikingly similar native-like folding intermediate across the three mutants, in which strand ß(1) is totally unstructured and more than half of the major hydrophobic core residues are highly solvent exposed. Results from extensive docking simulations show that the ability of the intermediates to dimerize is largely driven by strand ß(1) and is consistent with the in vitro aggregation behavior reported for the corresponding mutants. They further suggest that residues 44 and 53, which are key players in the nucleation-condensation mechanism of folding, are also important triggers of the aggregation process.

Identifying critical residues in protein folding: Insights from phi-value and P(fold) analysis.

We apply a simulational proxy of the phi-value analysis and perform extensive mutagenesis experiments to identify the nucleating residues in the folding "reactions" of two small lattice Go polymers with different native geometries. Our findings show that for the more complex native fold (i.e., the one that is rich in nonlocal, long-range bonds), mutation of the residues that form the folding nucleus leads to a considerably larger increase in the folding time than the corresponding mutations in the geometry that is predominantly local. These results are compared to data obtained from an accurate analysis based on the reaction coordinate folding probability P(fold) and on structural clustering methods. Our study reveals a complex picture of the transition state ensemble. For both protein models, the transition state ensemble is rather heterogeneous and splits up into structurally different populations. For the more complex geometry the identified subpopulations are actually structurally disjoint. For the less complex native geometry we found a broad transition state with microscopic heterogeneity. These findings suggest that the existence of multiple transition state structures may be linked to the geometric complexity of the native fold. For both geometries, the identification of the folding nucleus via the P(fold) analysis agrees with the identification of the folding nucleus carried out with the phi-value analysis. For the most complex geometry, however, the applied methodologies give more consistent results than for the more local geometry. The study of the transition state structure reveals that the nucleus residues are not necessarily fully native in the transition state. Indeed, it is only for the more complex geometry that two of the five critical residues show a considerably high probability of having all its native bonds formed in the transition state. Therefore, one concludes that, in general, the phi-value correlates with the acceleration/deceleration of folding induced by mutation, rather than with the degree of nativeness of the transition state, and that the "traditional" interpretation of phi-values may provide a more realistic picture of the structure of the transition state only for more complex native geometries.

Native geometry and the dynamics of protein folding.

In this paper, we investigate the role of native geometry on the kinetics of protein folding based on simple lattice models and Monte Carlo simulations. Results obtained within the scope of the Miyazawa-Jernigan indicate the existence of two dynamical folding regimes depending on the protein chain length. For chains larger than 80 amino acids, the folding performance is sensitive to the native state's conformation. Smaller chains, with less than 80 amino acids, fold via two-state kinetics and exhibit a significant correlation between the contact order parameter and the logarithmic folding times. In particular, chains with N=48 amino acids were found to belong to two broad classes of folding, characterized by different cooperativity, depending on the contact order parameter. Preliminary results based on the Go model show that the effect of long-range contact interaction strength in the folding kinetics is largely dependent on the native state's geometry.

Folding and form: insights from lattice simulations.

Monte Carlo simulations of a Miyazawa-Jernigan lattice-polymer model indicate that, depending on the native structure's geometry, the model exhibits two broad classes of folding mechanisms for two-state folders. Folding to native structures of low contact order is driven by backbone distance and is characterized by a progressive accumulation of structure towards the native fold. By contrast, folding to high contact order targets is dominated by intermediate stage contacts not present in the native fold, yielding a more cooperative folding process.