Simulation, experiment, and evolution: Understanding nucleation in protein S6 folding

Hubner, Isaac A.; Oliveberg, Mikael; Shakhnovich, Eugene I.

doi:10.1073/pnas.0401672101

Cited by 57 publications

(77 citation statements)

References 41 publications

(73 reference statements)

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“…O ur current understanding of the protein-folding nucleus is based on a synthesis of results from simulation (1) and experimental mapping of site-specific contacts in the transitionstate ensemble by protein engineering (2,3). From these results it is apparent that the free-energy landscape controlling the folding process is highly evolved, with few traps and a characteristic bias toward native contacts (4,5).…”

Section: Circular Permutation ͉ -Values ͉ Transition Statementioning

confidence: 99%

Identification of the minimal protein-folding nucleus through loop-entropy perturbations

Lindberg

Haglund

Hubner

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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To explore the plasticity and structural constraints of the proteinfolding nucleus we have constructed through circular permutation four topological variants of the ribosomal protein S6. In effect, these topological variants represent entropy mutants with maintained spatial contacts. The proteins were characterized at two complementary levels of detail: by -value analysis estimating the extent of contact formation in the transition-state ensemble and by Hammond analysis measuring the site-specific growth of the folding nucleus. The results show that, although the loop-entropy alterations markedly influence the appearance and structural location of the folding nucleus, it retains a common motif of one helix docking against two strands. This nucleation motif is built around a shared subset of side chains in the center of the hydrophobic core but extends in different directions of the S6 structure following the permutant-specific differences in local loop entropies. The adjustment of the critical folding nucleus to alterations in loop entropies is reflected by a direct correlation between the -value change and the accompanying change in local sequence separation.O ur current understanding of the protein-folding nucleus is based on a synthesis of results from simulation (1) and experimental mapping of site-specific contacts in the transitionstate ensemble by protein engineering (2, 3). From these results it is apparent that the free-energy landscape controlling the folding process is highly evolved, with few traps and a characteristic bias toward native contacts (4, 5). Consistent with the general insensitivity of the transition-state structure to point mutation and the remarkable success of reproducing experimental data with simplistic Go models, the prediction from such biased landscapes is that the sequence of folding events is largely determined by the topology of the native structure (1, 4, 6, 7). One intriguing possibility is that folding follows a trajectory of the lowest successive loop-entropy cost (8, 9). To directly test this idea we have analyzed the folding behavior of four S6 variants in which the loop-entropy cost of forming pairwise contacts has been systematically altered through circular permutation (10-12). The results show that the -value distribution defining the S6 nucleus is plastic and responds to circular permutation in a systematic manner. Contacts are recruited in directions with decreased sequence separation, and contacts are lost at the entropically penalized regions of the backbone incisions. Even so, the critical nuclei of the permuted proteins share a minimal two-strand-helix motif with variable but overlapping composition of secondary-structure elements. Moreover, it is apparent that a specific number of side-chain contacts are required to turn the folding free energy profile downhill, and that the dimension of this cluster matches the size of the smallest cooperatively folding proteins. Results and DiscussionPronounced Changes of the -Value Distribution upon Circular Permutation. Ch...

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Section: Circular Permutation ͉ -Values ͉ Transition Statementioning

confidence: 99%

Identification of the minimal protein-folding nucleus through loop-entropy perturbations

Lindberg

Haglund

Hubner

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…The results suggest that the envelop of residues that normally protects the hydrophobic interior of the SOD monomer is largely unfolded in the transition-state ensemble. Although the precise nature of this outer boundary of the transition-state structure is intrinsically difficult to pin down (37,38), it can be safely concluded that the side chains with low values consolidate mainly on the downhill side of the folding barrier. Vice versa, these side chains define the regions of the SOD structure that ruptures early in the unfolding process, i.e., what is missing at the top has been lost on the way up.…”

Section: The Aposod Monomer Folds Cooperatively According To a Two-statementioning

confidence: 99%

Folding of Cu/Zn superoxide dismutase suggests structural hotspots for gain of neurotoxic function in ALS: Parallels to precursors in amyloid disease

Nordlund

Oliveberg

2006

Proc. Natl. Acad. Sci. U.S.A.

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131

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Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease linked to misfolding of the ubiquitous enzyme Cu͞Zn superoxide dismutase (SOD). In contrast to other protein-misfolding disorders with similar neuropathogenesis, ALS is not always associated with the in vivo deposition of protein aggregates. Thus, under the assumption that all protein-misfolding disorders share at primary level a similar disease mechanism, ALS constitutes an interesting disease model for identifying the yet-mysterious precursor states from which the cytotoxic pathway emerges. In this study, we have mapped out the conformational repertoire of the apoSOD monomer through analysis of its folding behavior. The results allow us to target the regions of the SOD structure that are most susceptible to unfolding locally under physiological conditions, leading to the exposure of structurally promiscuous interfaces that are normally hidden in the protein's interior. The structure of this putative ALS precursor is strikingly similar to those implicated in amyloid disease.neurodegenerative disease ͉ protein folding ͉ transition state A n increasing number of severe neurodegenerative disorders have been linked to misfolding and subsequent aggregation of proteins (1). In some cases the protein aggregates deposit in the form of highly ordered amyloid fibrils (2), whereas in other cases they appear more granular or even amorphous (3). Even so, the main cause of neurotoxic function seems not to be the macroscopically deposited protein itself but rather a mysterious precursor species in the aggregation pathway (1, 4-6). In this perspective, it is notable that there are also disease cases where in vivo depositions of the disease-associated proteins are hard to find and sometimes are not even observable. Characteristic examples are found among the sporadic cases of CreuzfeldtJacob disease and amyotropic lateral sclerosis (ALS) (7). It is further apparent that the ALS-associated enzyme Cu͞Zn superoxide dismutase (SOD) has relatively low propensities to aggregate in vitro: SOD can be produced and analyzed by standard protocols despite being destabilized to the extent that it is half unfolded in physiological buffer at 25°C (8). For comparison there are few nondisease proteins that would resist aggregation under these conditions. The question then arises, do the disease cases with and without in vivo deposition of protein aggregates share at the primary level the same toxicity mechanism? We have previously observed that ALS-provoking SOD mutations produce a common shift of the folding equilibrium toward unfolded and partly folded monomers, implicating that the noxious side reaction emerges from the early folding events. The observation is in full agreement with a series of reports pointing at the immature apoSOD monomer as precursor for cytotoxicity (8)(9)(10)(11)(12)(13)(14)(15). Accompanying the shift of the folding equilibrium, the ALS-associated SOD mutations indicate a quantitative relation between decreased protein stability, net charge, and diseas...

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“…Just like the -value analysis in experiments with real-world proteins, the folding probability analysis method has been extensively used to investigate the TSE in computer simulations of protein folding. 13,[34][35][36][37] The P fold method has one caveat though: it is computationally costly. 38 Here we perform extensive Monte Carlo simulations of a simple lattice protein model to construct and compare two ensembles of conformations representative of the TSE.…”

Section: Introductionmentioning

confidence: 99%

The protein folding transition state: Insights from kinetics and thermodynamics

Travasso

Faísca

Rey

2010

The Journal of Chemical Physics

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We perform extensive lattice Monte Carlo simulations of protein folding to construct and compare the equilibrium and the kinetic transition state ensembles of a model protein that folds to the native state with two-state kinetics. The kinetic definition of the transition state is based on the folding probability analysis method, and therefore on the selection of conformations with 0.4Ͻ P fold Ͻ 0.6, while for the equilibrium characterization we consider conformations for which the evaluated values of several reaction coordinates correspond to the maximum of the free energy measured as a function of those reaction coordinates. Our results reveal a high degree of structural similarity between the ensembles determined by the two methods. However, the folding probability distribution of the conformations belonging to our definition of the equilibrium transition state ͑0.2Ͻ P fold Ͻ 0.8͒ is broader than that displayed by the kinetic transition state.

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Simulation, experiment, and evolution: Understanding nucleation in protein S6 folding

Cited by 57 publications

References 41 publications

Identification of the minimal protein-folding nucleus through loop-entropy perturbations

Identification of the minimal protein-folding nucleus through loop-entropy perturbations

Folding of Cu/Zn superoxide dismutase suggests structural hotspots for gain of neurotoxic function in ALS: Parallels to precursors in amyloid disease

The protein folding transition state: Insights from kinetics and thermodynamics

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