Landscape approaches for determining the ensemble of folding transition states: Success and failure hinge on the degree of frustration

100

A microscopic study of functional disorder-order folding transitions coupled to binding is performed for the p27 protein, which derives a kinetic advantage from the intrinsically disordered unbound form on binding with the phosphorylated cyclin A-cyclindependent kinase 2 (Cdk2) complex. Hierarchy of structural loss during p27 coupled unfolding and unbinding is simulated by using high-temperature Monte Carlo simulations initiated from the crystal structure of the tertiary complex. Subsequent determination of the transition-state ensemble and the proposed atomic picture of the folding mechanism coupled to binding provide a microscopic rationale that reconciles the initiation recruitment of p27 at the cyclin A docking site with the kinetic benefit for a disordered ␣-helix in the unbound form of p27. The emerging structural polarization in the ensemble of unfolding͞unbinding trajectories and in the computationally determined transition-state ensemble is not determined by the intrinsic folding preferences of p27 but rather is attributed to the topological requirements of the native intermolecular interface to order ␤-hairpin and ␤-strand of p27 that could be critical for nucleating rapid folding transition coupled to binding. In agreement with the experimental data, the disorderorder folding transition for p27 is largely determined by the functional requirement to form a specific intermolecular interface that ultimately dictates the folding mechanism and overwhelms any local folding preferences for creating a stable ␣-helix in the p27 structure before overcoming the major free energy barrier.

mentioning

confidence: 50%

Simulating disorder–order transitions in molecular recognition of unstructured proteins: Where folding meets binding

Verkhivker

Bouzida²,

Gehlhaar³

et al. 2003

100

Section: A Minimalist Model For Water-mediated Interactionmentioning

confidence: 99%

“…Again, although no nonnative attractive interactions are included in these unfrustrated models, what appears to be unrealistic, recent theoretical and experimental results support their applicability. In addition to the experimental citations earlier in the introduction, several recent theoretical͞simulational approaches have shown the geometrical features of the folding mechanism are robust to energetic frustration as long as it is small relative to the bias toward the native configuration (34,35).Therefore, even when neglecting the effects of energetic frustration, these Go models have been shown to reproduce nearly all-global geometrical features of the transition state ensemble and intermediates of the real, fast folding proteins (16,20,22). The new interactions, however, include not only the standard minimum for the direct contacts between residues, but…”

mentioning

confidence: 99%

Protein folding mediated by solvation: Water expulsion and formation of the hydrophobic core occur after the structural collapse

Cheung

Garcı́a

Onuchic

2002

466

404

The interplay between structure-search of the native structure and desolvation in protein folding has been explored using a minimalist model. These results support a folding mechanism where most of the structural formation of the protein is achieved before water is expelled from the hydrophobic core. This view integrates water expulsion effects into the funnel energy landscape theory of protein folding. Comparisons to experimental results are shown for the SH3 protein. After the folding transition, a near-native intermediate with partially solvated hydrophobic core is found. This transition is followed by a final step that cooperatively squeezes out water molecules from the partially hydrated protein core.T he energy landscape theory and the funnel concept combined with a new generation of experiments provide a new framework for understanding the protein folding problem. According to this ''new view,'' the folding landscape for proteins (at least for small fast folding proteins) resembles a partially rough funnel riddled with traps where the protein can transiently reside (1-8). Recent theoretical and experimental evidence fully support this picture and suggests that, because the energetic roughness of the landscape is sufficiently small (low level of energetic frustration), the global characteristics of the structural heterogeneity observed in the transition state ensemble for two-state fast folding proteins are most strongly determined by the native state topology. For this reason, theoretical energetically unfrustrated models have been shown to reproduce nearly all experimental results for the global geometrical features of the transition state ensemble of a large number of real proteins, which are two-state or three-state folders, on whose native structures they are based (9-24).Some concerns have been raised, however, about the applicability of these models. For example, solvent effects are incorporated implicitly into the energetic interactions between residues. Such implicit equilibrium models for solvation are unable to capture effects such as desolvation of the hydrophobic core; therefore, understanding the potential of mean force for the interaction between solutes in an aqueous solvent attracts the attention of many research groups (25-30). The potential of mean force (PMF) between two methane-like particles exhibit two minima-a contact minimum at the van der Waals distance between the particles and a solvent-separated minimum. The two minima are separated by a desolvation barrier with a width of approximately one water molecule diameter. Recently, Hillson et al. (31,32) advertised the importance of the desolvation effect by taking into account the process of expelling a water molecule in a pairwise interaction between hydrophobic particles. Assuming that the time scale for water penetration and escape are faster than the contact formation in proteins (33), the shape of this potential energy function implies that there are free-energy costs for protein (de)solvation (27). The time scale separation ass...

“…Identifying a protein's TS can provide crucial insight into its folding mechanism. The TS ensemble (TSE) is difficult to determine both experimentally and computationally (27,28). Experimentally, -value analysis is used to infer the structure of the TSE (29).…”

Section: Tethering Alters Thermodynamic Stability and Protein Foldingmentioning

confidence: 99%

Effects of surface tethering on protein folding mechanisms

Friedel¹,

Baumketner

Shea

2006

100

113

The folding mechanisms of proteins are increasingly being probed through single-molecule experiments in which the protein is immobilized on a surface. Nevertheless, a clear understanding of how the surface might affect folding, and whether or not it changes folding from its bulk behavior, is lacking. In this work, we use molecular dynamics simulations of a model ␤-barrel protein tethered to a surface to systematically investigate how the surface impacts folding. In the bulk, this protein folds in a three-state manner through a compact intermediate state, and its transition state (TS) has a well formed hydrophobic core. Upon tethering, we find that folding rates and stability are impacted differently by the surface, with dependencies on both the length and location of the tether. Significant changes in folding times are observed for tether points that do not alter the folding temperature. Tethering also locally enhances the formation of structure for residues proximal to the tether point. We find that neither the folding mechanism nor the TS of this protein are altered if the tether is in a fully structured or completely unstructured region of the TS. By contrast, tethering in a partially structured region of the TS leads to dramatic changes. For one such tether point, the intermediate present in bulk folding is eliminated, leading to a two-state folding process with a heterogeneous, highly unstructured TS ensemble. These results have implications for both the design of single-molecule experiments and biotechnological applications of tethered proteins. molecular dynamics simulations ͉ protein-surface interactions ͉ single-molecule spectroscopy S ingle-molecule spectroscopy has recently emerged as a powerful technique for watching individual proteins fold (1-3). By attaching donor and acceptor dyes to key residues of a protein, fluorescence resonance energy transfer (FRET), already successful in ensemble folding experiments (4), can be used as a distance probe to monitor individual folding pathways. There are several different experimental techniques for doing FRET-based single-molecule experiments on proteins, each with distinct advantages and challenges. Bulk experiments use a focusing laser beam that monitors folding as proteins diffuse freely through the area illuminated by the laser (5, 6). Although this approach is advantageous in that the protein is allowed to fold in a relatively nondisrupted manner, solution experiments are diffusion limited and cannot examine slower (տ10 ms) phenomena (2). Proteins enclosed in surface-tethered vesicles allow observations on a more spatially localized scale than the bulk but do not allow for the rapid exchange of buffer conditions or the use of extreme denaturing environments (7). By immobilizing them directly on a surface (8), proteins can be observed in both a spatially localized region and over longer time scales than those accessible in diffusion-limited experiments. Despite these advantages, the folding behavior of surface-tethered proteins also may be influenced by ...