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Chemphyschem : a European Journal of... Mar 2020We present a computational study on the folding and aggregation of proteins in an aqueous environment, as a function of its concentration. We show how the increase of...
We present a computational study on the folding and aggregation of proteins in an aqueous environment, as a function of its concentration. We show how the increase of the concentration of individual protein species can induce a partial unfolding of the native conformation without the occurrence of aggregates. A further increment of the protein concentration results in the complete loss of the folded structures and induces the formation of protein aggregates. We discuss the effect of the protein interface on the water fluctuations in the protein hydration shell and their relevance in the protein-protein interaction.
Topics: Algorithms; Hydrophobic and Hydrophilic Interactions; Molecular Dynamics Simulation; Protein Aggregates; Protein Conformation; Protein Unfolding; Proteins; Thermodynamics
PubMed: 31721405
DOI: 10.1002/cphc.201900904 -
Current Opinion in Biotechnology Aug 1996Protein unfolding occurs when the balance of forces between the protein's interaction with itself and the protein's interaction with its environment is disrupted. The... (Review)
Review
Protein unfolding occurs when the balance of forces between the protein's interaction with itself and the protein's interaction with its environment is disrupted. The disruption of this balance of forces may be as simple as a perturbance of the normal water structure around the protein. A decrease in the normal water-water interaction will result in an increase in the relative interaction of water with the protein. An increase in the number of interactions between water and the protein may initiate a protein's unfolding. This model for protein unfolding is supported by a range of recent experimental and computational data.
Topics: Computer Simulation; Models, Molecular; Protein Denaturation; Protein Folding; Proteins; Solvents; Urea; Water
PubMed: 8768902
DOI: 10.1016/s0958-1669(96)80119-4 -
Advanced Science (Weinheim,... Feb 2022Protein-based hydrogels have attracted great attention due to their excellent biocompatible properties, but often suffer from weak mechanical strength. Conventional...
Protein-based hydrogels have attracted great attention due to their excellent biocompatible properties, but often suffer from weak mechanical strength. Conventional strengthening strategies for protein-based hydrogels are to introduce nanoparticles or synthetic polymers for improving their mechanical strength, but often compromise their biocompatibility. Here, a new, general, protein unfolding-chemical coupling (PNC) strategy is developed to fabricate pure protein hydrogels without any additives to achieve both high mechanical strength and excellent cell biocompatibility. This PNC strategy combines thermal-induced protein unfolding/gelation to form a physically-crosslinked network and a -NH2/-COOH coupling reaction to generate a chemicallycrosslinked network. Using bovine serum albumin (BSA) as a globular protein, PNC-BSA hydrogels show macroscopic transparency, high stability, high mechanical properties (compressive/tensile strength of 115/0.43 MPa), fast stiffness/toughness recovery of 85%/91% at room temperature, good fatigue resistance, and low cell cytotoxicity and red blood cell hemolysis. More importantly, the PNC strategy can be not only generally applied to silk fibroin, ovalbumin, and milk albumin protein to form different, high strength protein hydrogels, but also modified with PEDOT/PSS nanoparticles as strain sensors and fluorescent fillers as color sensors. This work demonstrates a new, universal, PNC method to prepare high strength, multi-functional, pure protein hydrogels beyond a few available today.
Topics: Fibroins; Hydrogels; Polymers; Protein Unfolding; Serum Albumin, Bovine
PubMed: 34939355
DOI: 10.1002/advs.202102557 -
The Journal of Physical Chemistry. B Apr 2023Protein stability is important in many areas of life sciences. Thermal protein unfolding is investigated extensively with various spectroscopic techniques. The...
Protein stability is important in many areas of life sciences. Thermal protein unfolding is investigated extensively with various spectroscopic techniques. The extraction of thermodynamic properties from these measurements requires the application of models. Differential scanning calorimetry (DSC) is less common, but is unique as it measures directly a thermodynamic property, that is, the heat capacity (). The analysis of () is usually performed with the chemical equilibrium two-state model. This is not necessary and leads to incorrect thermodynamic consequences. Here we demonstrate a straightforward model-independent evaluation of heat capacity experiments in terms of protein unfolding enthalpy Δ(), entropy Δ(), and free energy Δ()). This now allows the comparison of the experimental thermodynamic data with the predictions of different models. We critically examined the standard chemical equilibrium two-state model, which predicts a positive free energy for the native protein, and diverges distinctly from the experimental temperature profiles. We propose two new models which are equally applicable to spectroscopy and calorimetry. The Θ()-weighted chemical equilibrium model and the statistical-mechanical two-state model provide excellent fits of the experimental data. They predict sigmoidal temperature profiles for enthalpy and entropy, and a trapezoidal temperature profile for the free energy. This is illustrated with experimental examples for heat and cold denaturation of lysozyme and β-lactoglobulin. We then show that the free energy is not a good criterion to judge protein stability. More useful parameters are discussed, including protein cooperativity. The new parameters are embedded in a well-defined thermodynamic context and are amenable to molecular dynamics calculations.
Topics: Hot Temperature; Protein Denaturation; Proteins; Thermodynamics; Cold Temperature; Protein Unfolding; Calorimetry, Differential Scanning; Protein Folding
PubMed: 37040567
DOI: 10.1021/acs.jpcb.3c00882 -
Chembiochem : a European Journal of... Aug 2023This review aims to analyse the role of solution nuclear magnetic resonance spectroscopy in pressure-induced in vitro studies of protein unfolding. Although this... (Review)
Review
This review aims to analyse the role of solution nuclear magnetic resonance spectroscopy in pressure-induced in vitro studies of protein unfolding. Although this transition has been neglected for many years because of technical difficulties, it provides important information about the forces that keep protein structure together. We first analyse what pressure unfolding is, then provide a critical overview of how NMR spectroscopy has contributed to the field and evaluate the observables used in these studies. Finally, we discuss the commonalities and differences between pressure-, cold- and heat-induced unfolding. We conclude that, despite specific peculiarities, in both cold and pressure denaturation the important contribution of the state of hydration of nonpolar side chains is a major factor that determines the pressure dependence of the conformational stability of proteins.
Topics: Protein Denaturation; Proteins; Magnetic Resonance Spectroscopy; Protein Unfolding; Protein Conformation; Thermodynamics; Protein Folding; Cold Temperature
PubMed: 37154795
DOI: 10.1002/cbic.202300164 -
ACS Nano Jul 2022Globular folded proteins are versatile nanoscale building blocks to create biomaterials with mechanical robustness and inherent biological functionality due to their...
Globular folded proteins are versatile nanoscale building blocks to create biomaterials with mechanical robustness and inherent biological functionality due to their specific and well-defined folded structures. Modulating the nanoscale unfolding of protein building blocks during network formation ( protein unfolding) provides potent opportunities to control the protein network structure and mechanics. Here, we control protein unfolding during the formation of hydrogels constructed from chemically cross-linked maltose binding protein using ligand binding and the addition of cosolutes to modulate protein kinetic and thermodynamic stability. Bulk shear rheology characterizes the storage moduli of the bound and unbound protein hydrogels and reveals a correlation between network rigidity, characterized as an increase in the storage modulus, and protein thermodynamic stability. Furthermore, analysis of the network relaxation behavior identifies a crossover from an unfolding dominated regime to an entanglement dominated regime. Control of protein unfolding and entanglement provides an important route to finely tune the architecture, mechanics, and dynamic relaxation of protein hydrogels. Such predictive control will be advantageous for future smart biomaterials for applications which require responsive and dynamic modulation of mechanical properties and biological function.
Topics: Hydrogels; Biocompatible Materials; Rheology; Proteins; Protein Unfolding
PubMed: 35731007
DOI: 10.1021/acsnano.2c02369 -
Langmuir : the ACS Journal of Surfaces... Jul 2018Dynamic modulation of lipid membrane curvature can be achieved by a number of peripheral protein binding mechanisms such as hydrophobic insertion of amphipathic helices...
Dynamic modulation of lipid membrane curvature can be achieved by a number of peripheral protein binding mechanisms such as hydrophobic insertion of amphipathic helices and membrane scaffolding. Recently, an alternative mechanism was proposed in which crowding of peripherally bound proteins induces membrane curvature through steric pressure generated by lateral collisions. This effect was enhanced using intrinsically disordered proteins that possess high hydrodynamic radii, prompting us to explore whether membrane bending can be triggered by the folding-unfolding transition of surface-bound proteins. We utilized histidine-tagged human serum albumin bound to Ni-NTA-DGS containing liposomes as our model system to test this hypothesis. We found that reduction of the disulfide bonds in the protein resulted in unfolding of HSA, which subsequently led to membrane tubule formation. The frequency of tubule formation was found to be significantly higher when the proteins were unfolded while being localized to a phase-separated domain as opposed to randomly distributed in fluid phase liposomes, indicating that the steric pressure generated from protein unfolding can drive membrane deformation. Our results are critical for the design of peripheral membrane protein-immobilization strategies and open new avenues for exploring mechanisms of membrane bending driven by conformational changes of peripheral membrane proteins.
Topics: Cell Membrane Structures; Humans; Hydrophobic and Hydrophilic Interactions; Membrane Proteins; Protein Binding; Protein Structure, Secondary; Protein Unfolding; Serum Albumin
PubMed: 29925237
DOI: 10.1021/acs.langmuir.8b01136 -
Biophysical Chemistry Feb 2018Knowledge of protein stability is of utmost importance in various fields of biotechnology. Protein stability can be assessed in solution by increasing the concentration...
Knowledge of protein stability is of utmost importance in various fields of biotechnology. Protein stability can be assessed in solution by increasing the concentration of denaturant and recording the structural changes with spectroscopic or thermodynamic methods. The standard interpretation of the experimental data is to assume a 2-state equilibrium between completely folded and completely unfolded protein molecules. Here we propose a cooperative model based on the statistical-mechanical Zimm-Bragg theory. In this model protein unfolding is driven by the weak binding of a rather small number of denaturant molecules, inducing the cooperative unfolding with multiple dynamic intermediates. The modified Zimm-Bragg theory is applied to published thermodynamic and spectroscopic data leading to the following conclusions. (i) The binding constant K is correlated with the midpoint concentration, c, of the unfolding reaction according to c≅1/K. The better the binding of denaturant the lower is the concentration to achieve unfolding. (ii) The binding constant K agrees with direct thermodynamic measurements. A rather small number of bound denaturants suffices to induce the cooperative unfolding of the whole protein. (iii) Chemical unfolding occurs in the concentration range Δc=c-c. The theory predicts the unfolding energy per amino acid residue as g=RTK(c-c). The Gibbs free energy of an osmotic gradient of the same size is ΔG=-RTln(c/c). In all examples investigated ΔG exactly balances the unfolding energy g. The total unfolding energy is thus close to zero. (iv) Protein cooperativity in chemical unfolding is rather low with cooperativity parameters σ≥3x10. As a consequence, the theory predicts a dynamic mixture of conformations during the unfolding reaction. The probabilities of individual conformations are easily accessible via the partition function Z(c,σ).
Topics: Bacterial Proteins; Chymotrypsin; Hydrochloric Acid; Models, Statistical; Muramidase; Phosphoenolpyruvate Sugar Phosphotransferase System; Protein Denaturation; Protein Unfolding; Thermodynamics; Urea
PubMed: 29232602
DOI: 10.1016/j.bpc.2017.12.001 -
Biomolecules Sep 2020In this work, we investigate the role of folding/unfolding equilibrium in protein aggregation and formation of a gel network. Near the neutral pH and at a low buffer...
In this work, we investigate the role of folding/unfolding equilibrium in protein aggregation and formation of a gel network. Near the neutral pH and at a low buffer ionic strength, the formation of the gel network around unfolding conditions prevents investigations of protein aggregation. In this study, by deploying the fact that in lysozyme solutions the time of folding/unfolding is much shorter than the characteristic time of gelation, we have prevented gelation by rapidly heating the solution up to the unfolding temperature (~80 °C) for a short time (~30 min.) followed by fast cooling to the room temperature. Dynamic light scattering measurements show that if the gelation is prevented, nanosized irreversible aggregates (about 10-15 nm radius) form over a time scale of 10 days. These small aggregates persist and aggregate further into larger aggregates over several weeks. If gelation is not prevented, the nanosized aggregates become the building blocks for the gel network and define its mesh length scale. These results support our previously published conclusion on the nature of mesoscopic aggregates commonly observed in solutions of lysozyme, namely that aggregates do not form from lysozyme monomers in their native folded state. Only with the emergence of a small fraction of unfolded proteins molecules will the aggregates start to appear and grow.
Topics: Dynamic Light Scattering; Gels; Hot Temperature; Muramidase; Protein Aggregates; Protein Unfolding; Solutions
PubMed: 32887233
DOI: 10.3390/biom10091262 -
The Journal of Physical Chemistry. B Feb 2021Deviations from linearity in the dependence of the logarithm of protein unfolding rates, log (), as a function of mechanical force, , measurable in single molecule...
Deviations from linearity in the dependence of the logarithm of protein unfolding rates, log (), as a function of mechanical force, , measurable in single molecule experiments, can arise for many reasons. In particular, upward curvature in log () as a function of implies that the underlying energy landscape must be multidimensional with the possibility that unfolding ensues by parallel pathways. Here, simulations using the SOP-SC model of a wild type β-sandwich protein and several mutants, with immunoglobulin folds, show upward curvature in the unfolding kinetics. There are substantial changes in the structures of the transition state ensembles as the force is increased, signaling a switch in the unfolding pathways. Our results, when combined with previous theoretical and experimental studies, show that parallel unfolding of structurally unrelated single domain proteins can be determined from the dependence of log () as a function of force (or log [C] where [C] is the denaturant concentration).
Topics: Kinetics; Protein Denaturation; Protein Folding; Protein Unfolding; Proteins
PubMed: 33565314
DOI: 10.1021/acs.jpcb.0c11308