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The Biochemical Journal Sep 2016Protein folding research stalled for decades because conventional experiments indicated that proteins fold slowly and in single strokes, whereas theory predicted a... (Review)
Review
Protein folding research stalled for decades because conventional experiments indicated that proteins fold slowly and in single strokes, whereas theory predicted a complex interplay between dynamics and energetics resulting in myriad microscopic pathways. Ultrafast kinetic methods turned the field upside down by providing the means to probe fundamental aspects of folding, test theoretical predictions and benchmark simulations. Accordingly, experimentalists could measure the timescales for all relevant folding motions, determine the folding speed limit and confirm that folding barriers are entropic bottlenecks. Moreover, a catalogue of proteins that fold extremely fast (microseconds) could be identified. Such fast-folding proteins cross shallow free energy barriers or fold downhill, and thus unfold with minimal co-operativity (gradually). A new generation of thermodynamic methods has exploited this property to map folding landscapes, interaction networks and mechanisms at nearly atomic resolution. In parallel, modern molecular dynamics simulations have finally reached the timescales required to watch fast-folding proteins fold and unfold in silico All of these findings have buttressed the fundamentals of protein folding predicted by theory, and are now offering the first glimpses at the underlying mechanisms. Fast folding appears to also have functional implications as recent results connect downhill folding with intrinsically disordered proteins, their complex binding modes and ability to moonlight. These connections suggest that the coupling between downhill (un)folding and binding enables such protein domains to operate analogically as conformational rheostats.
Topics: Protein Folding; Proteins; Thermodynamics
PubMed: 27574021
DOI: 10.1042/BCJ20160107 -
Molecular Biology of the Cell Nov 2017Protein folding in the cell was originally assumed to be a spontaneous process, based on Anfinsen's discovery that purified proteins can fold on their own after removal...
Protein folding in the cell was originally assumed to be a spontaneous process, based on Anfinsen's discovery that purified proteins can fold on their own after removal from denaturant. Consequently cell biologists showed little interest in the protein folding process. This changed only in the mid and late 1980s, when the chaperone story began to unfold. As a result, we now know that in vivo, protein folding requires assistance by a complex machinery of molecular chaperones. To ensure efficient folding, members of different chaperone classes receive the nascent protein chain emerging from the ribosome and guide it along an ordered pathway toward the native state. I was fortunate to contribute to these developments early on. In this short essay, I will describe some of the critical steps leading to the current concept of protein folding as a highly organized cellular process.
Topics: Mitochondria; Models, Molecular; Molecular Chaperones; Protein Folding
PubMed: 29084909
DOI: 10.1091/mbc.E17-07-0480 -
International Journal of Molecular... Nov 2020Disordered proteins often act as interaction hubs in cellular pathways, via the specific recognition of a distinguished set of partners. While disordered regions can... (Review)
Review
Disordered proteins often act as interaction hubs in cellular pathways, via the specific recognition of a distinguished set of partners. While disordered regions can adopt a well-defined conformation upon binding, the coupled folding to binding model does not explain how interaction versatility is achieved. Here, I present a classification scheme for the binding modes of disordered protein regions, based on their conformational heterogeneity in the bound state. Binding modes are defined as (i) leading to a well-defined bound state, (ii) leading to a disordered bound state and (iii) when the degree of disorder in the bound state may vary with the partner or cellular conditions. Fuzzy binding includes polymorphic bound structures, conditional folding and dynamic binding. This classification scheme describes the structural continuum of complexes involving disordered regions as well as their context-dependent interaction behaviors.
Topics: Intrinsically Disordered Proteins; Models, Molecular; Protein Folding
PubMed: 33207556
DOI: 10.3390/ijms21228615 -
Biotechnology Advances 2020In the biological milieu of a cell, soluble crowding molecules and rigid confined environments strongly influence whether the protein is properly folded, intrinsically... (Review)
Review
In the biological milieu of a cell, soluble crowding molecules and rigid confined environments strongly influence whether the protein is properly folded, intrinsically disordered proteins assemble into distinct phases, or a denatured or aggregated protein species is favored. Such crowding and confinement factors act to exclude solvent volume from the protein molecules, resulting in an increased local protein concentration and decreased protein entropy. A protein's structure is inherently tied to its function. Examples of processes where crowding and confinement may strongly influence protein function include transmembrane protein dimerization, enzymatic activity, assembly of supramolecular structures (e.g., microtubules), nuclear condensates containing transcriptional machinery, protein aggregation in the contexts of disease and protein therapeutics. Historically, most protein structures have been determined from pure, dilute protein solutions or pure crystals. However, these are not the environments in which these proteins function. Thus, there has been an increased emphasis on analyzing protein structure and dynamics in more "in vivo-like" environments. Complex in vitro models using hydrogel scaffolds to study proteins may better mimic features of the in vivo environment. Therefore, analytical techniques need to be optimized for real-time analysis of proteins within hydrogel scaffolds.
Topics: Hydrogels; Protein Aggregates; Protein Folding; Proteins
PubMed: 32512220
DOI: 10.1016/j.biotechadv.2020.107573 -
The Journal of Biological Chemistry Nov 2021The major classes of molecular chaperones have highly variable sequences, sizes, and shapes, yet they all bind to unfolded proteins, limit their aggregation, and assist... (Review)
Review
The major classes of molecular chaperones have highly variable sequences, sizes, and shapes, yet they all bind to unfolded proteins, limit their aggregation, and assist in their folding. Despite the central importance of this process to protein homeostasis, it has not been clear exactly how chaperones guide this process or whether the diverse families of chaperones use similar mechanisms. For the first time, recent advances in NMR spectroscopy have enabled detailed studies of how unfolded, "client" proteins interact with both ATP-dependent and ATP-independent classes of chaperones. Here, we review examples from four distinct chaperones, Spy, Trigger Factor, DnaK, and HscA-HscB, highlighting the similarities and differences between their mechanisms. One striking similarity is that the chaperones all bind weakly to their clients, such that the chaperone-client interactions are readily outcompeted by stronger, intra- and intermolecular contacts in the folded state. Thus, the relatively weak affinity of these interactions seems to provide directionality to the folding process. However, there are also key differences, especially in the details of how the chaperones release clients and how ATP cycling impacts that process. For example, Spy releases clients in a largely folded state, while clients seem to be unfolded upon release from Trigger Factor or DnaK. Together, these studies are beginning to uncover the similarities and differences in how chaperones use weak interactions to guide protein folding.
Topics: Humans; Molecular Chaperones; Protein Binding; Protein Conformation; Protein Folding
PubMed: 34624315
DOI: 10.1016/j.jbc.2021.101282 -
Molecules (Basel, Switzerland) Jul 2022Despite the recent advances in the prediction of protein structures by deep neutral networks, the elucidation of protein-folding mechanisms remains challenging. A... (Review)
Review
Despite the recent advances in the prediction of protein structures by deep neutral networks, the elucidation of protein-folding mechanisms remains challenging. A promising theory for describing protein folding is a coarse-grained statistical mechanical model called the Wako-Saitô-Muñoz-Eaton (WSME) model. The model can calculate the free-energy landscapes of proteins based on a three-dimensional structure with low computational complexity, thereby providing a comprehensive understanding of the folding pathways and the structure and stability of the intermediates and transition states involved in the folding reaction. In this review, we summarize previous and recent studies on protein folding and dynamics performed using the WSME model and discuss future challenges and prospects. The WSME model successfully predicted the folding mechanisms of small single-domain proteins and the effects of amino-acid substitutions on protein stability and folding in a manner that was consistent with experimental results. Furthermore, extended versions of the WSME model were applied to predict the folding mechanisms of multi-domain proteins and the conformational changes associated with protein function. Thus, the WSME model may contribute significantly to solving the protein-folding problem and is expected to be useful for predicting protein folding, stability, and dynamics in basic research and in industrial and medical applications.
Topics: Kinetics; Models, Molecular; Protein Folding; Protein Stability; Proteins; Thermodynamics
PubMed: 35889332
DOI: 10.3390/molecules27144460 -
Annual Review of Biophysics May 2021Cooperativity is a hallmark of protein folding, but the thermodynamic origins of cooperativity are difficult to quantify. Tandem repeat proteins provide a unique... (Review)
Review
Cooperativity is a hallmark of protein folding, but the thermodynamic origins of cooperativity are difficult to quantify. Tandem repeat proteins provide a unique experimental system to quantify cooperativity due to their internal symmetry and their tolerance of deletion, extension, and in some cases fragmentation into single repeats. Analysis of repeat proteins of different lengths with nearest-neighbor Ising models provides values for repeat folding ([Formula: see text]) and inter-repeat coupling (Δ). In this article, we review the architecture of repeat proteins and classify them in terms of Δ and Δ; this classification scheme groups repeat proteins according to their degree of cooperativity. We then present various statistical thermodynamic models, based on the 1D-Ising model, for analysis of different classes of repeat proteins. We use these models to analyze data for highly and moderately cooperative and noncooperative repeat proteins and relate their fitted parameters to overall structural features.
Topics: Models, Molecular; Protein Folding; Tandem Repeat Sequences; Thermodynamics
PubMed: 33606943
DOI: 10.1146/annurev-biophys-102220-083020 -
Progress in Biophysics and Molecular... Sep 2017In domain-swapping, two or more identical protein monomers exchange structural elements and fold into dimers or multimers whose units are structurally similar to the... (Review)
Review
In domain-swapping, two or more identical protein monomers exchange structural elements and fold into dimers or multimers whose units are structurally similar to the original monomer. Domain-swapping is of biotechnological interest because inhibiting domain-swapping can reduce disease-causing fibrillar protein aggregation. To achieve such inhibition, it is important to understand both the energetics that stabilize the domain-swapped structure and the protein dynamics that enable the swapping. Structure-based models (SBMs) encode the folded structure of the protein in their potential energy functions. SBMs have been successfully used to understand diverse aspects of monomer folding. Symmetrized SBMs model interactions between two identical protein chains using only intra-monomer interactions. Molecular dynamics simulations of such symmetrized SBMs have been used to correctly predict the domain-swapped structure and to understand the mechanism of domain-swapping. Here, we review such models and illustrate that monomer topology determines key aspects of domain-swapping. However, in some proteins, specifics of local energetic interactions modulate domain-swapping and these need to be added to the symmetrized SBMs. We then summarize some general principles of the mechanism of domain-swapping that emerge from the symmetrized SBM simulations. Finally, using our own results, we explore how symmetrized SBMs could be used to design domain-swapping in proteins.
Topics: Models, Molecular; Protein Domains; Protein Folding; Proteins
PubMed: 27867057
DOI: 10.1016/j.pbiomolbio.2016.09.013 -
Proceedings of the National Academy of... Nov 2014How do proteins fold, and why do they fold in that way? This Perspective integrates earlier and more recent advances over the 50-y history of the protein folding... (Review)
Review
How do proteins fold, and why do they fold in that way? This Perspective integrates earlier and more recent advances over the 50-y history of the protein folding problem, emphasizing unambiguously clear structural information. Experimental results show that, contrary to prior belief, proteins are multistate rather than two-state objects. They are composed of separately cooperative foldon building blocks that can be seen to repeatedly unfold and refold as units even under native conditions. Similarly, foldons are lost as units when proteins are destabilized to produce partially unfolded equilibrium molten globules. In kinetic folding, the inherently cooperative nature of foldons predisposes the thermally driven amino acid-level search to form an initial foldon and subsequent foldons in later assisted searches. The small size of foldon units, ∼ 20 residues, resolves the Levinthal time-scale search problem. These microscopic-level search processes can be identified with the disordered multitrack search envisioned in the "new view" model for protein folding. Emergent macroscopic foldon-foldon interactions then collectively provide the structural guidance and free energy bias for the ordered addition of foldons in a stepwise pathway that sequentially builds the native protein. These conclusions reconcile the seemingly opposed new view and defined pathway models; the two models account for different stages of the protein folding process. Additionally, these observations answer the "how" and the "why" questions. The protein folding pathway depends on the same foldon units and foldon-foldon interactions that construct the native structure.
Topics: Kinetics; Models, Chemical; Protein Folding
PubMed: 25326421
DOI: 10.1073/pnas.1411798111 -
The Journal of Physical Chemistry. B Oct 2019Cells of the vast majority of organisms are subject to temperature, pressure, pH, ionic strength, and other stresses. We discuss these effects in the light of protein... (Review)
Review
Cells of the vast majority of organisms are subject to temperature, pressure, pH, ionic strength, and other stresses. We discuss these effects in the light of protein folding and protein interactions , in complex environments, in cells, and . Protein phase diagrams provide a way of organizing different structural ensembles that occur under stress and how one can move among ensembles. Experiments that perturb biomolecules or in cells by stressing them have revealed much about the underlying forces that are competing to control protein stability, folding, and function. Two phenomena that emerge and serve to broadly classify effects of the cellular environment are crowding (mainly due to repulsive forces) and sticking (mainly due to attractive forces). The interior of cells is closely balanced between these emergent effects, and stress can tip the balance one way or the other. The free energy scale involved is small but significant on the scale of the "on/off switches" that control signaling in cells or of protein-protein association with a favorable function such as increased enzyme processivity. Quantitative tools from biophysical chemistry will play an important role in elucidating the world of crowding and sticking under stress.
Topics: Animals; Humans; Protein Binding; Protein Folding; Proteins; Stress, Physiological
PubMed: 31386813
DOI: 10.1021/acs.jpcb.9b05467