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Roland Winter - One of the best experts on this subject based on the ideXlab platform.
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Structural Basis for the Dissociation of Alpha-Synuclein Fibrils Triggered by Pressure Perturbation of the Hydrophobic Core
Biophysical Journal, 2017Co-Authors: Guilherme A P De Oliveira, Roland Winter, Mayra De A Marques, Yraima Cordeiro, Caroline Schuabb, Adolfo H Moraes, Hartmut Oschkinat, Debora Foguel, Mônica S. Freitas, Jerson L. SilvaAbstract:Parkinson's disease is a neurological disease in which aggregated forms of the alpha-synuclein (alpha-syn) protein are found. We used high hydrostatic Pressure (HHP) coupled with NMR spectroscopy to study the structure and dynamics of alpha-syn monomeric species released from fibrils. Different dynamic properties in the non-amyloid-beta component (NAC), which constitutes the Greek-key hydrophobic core, and in the acidic C-terminal region of the protein were identified by HHP NMR spectroscopy. In addition, solid-state NMR revealed subtle differences in the HHP-disturbed fibril core, providing clues to how these species contribute to seeding alpha-syn aggregation. These findings show how Pressure can populate so far undetected alpha-syn species, and they lay out a roadmap for fibril dissociation via pathways not previously observed using other approaches. Pressure perturbs the cavity-prone hydrophobic core of the fibrils by pushing water inward, thereby inducing the dissociation into monomers. Our study offers the molecular details of how hydrophobic interaction and the formation of water-excluded cavities jointly contribute to the assembly and stabilization of the fibrils. Understanding the molecular forces behind the formation of pathogenic fibrils uncovered by Pressure Perturbation will aid in the development of new therapeutics against Parkinson's disease.
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structural basis for the dissociation of α synuclein fibrils triggered by Pressure Perturbation of the hydrophobic core
Scientific Reports, 2016Co-Authors: Guilherme A P De Oliveira, Roland Winter, Mayra De A Marques, Carolina Cruzeirosilva, Yraima Cordeiro, Caroline Schuabb, Adolfo H Moraes, Hartmut Oschkinat, Debora FoguelAbstract:Parkinson's disease is a neurological disease in which aggregated forms of the α-synuclein (α-syn) protein are found. We used high hydrostatic Pressure (HHP) coupled with NMR spectroscopy to study the dissociation of α-syn fibril into monomers and evaluate their structural and dynamic properties. Different dynamic properties in the non-amyloid-β component (NAC), which constitutes the Greek-key hydrophobic core, and in the acidic C-terminal region of the protein were identified by HHP NMR spectroscopy. In addition, solid-state NMR revealed subtle differences in the HHP-disturbed fibril core, providing clues to how these species contribute to seeding α-syn aggregation. These findings show how Pressure can populate so far undetected α-syn species, and they lay out a roadmap for fibril dissociation via pathways not previously observed using other approaches. Pressure perturbs the cavity-prone hydrophobic core of the fibrils by pushing water inward, thereby inducing the dissociation into monomers. Our study offers the molecular details of how hydrophobic interaction and the formation of water-excluded cavities jointly contribute to the assembly and stabilization of the fibrils. Understanding the molecular forces behind the formation of pathogenic fibrils uncovered by Pressure Perturbation will aid in the development of new therapeutics against Parkinson's disease.
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Pressure Perturbation: A Prime Tool to Study Conformational Substates and Volume Fluctuations of Biomolecular Assemblies
Molecular Science of Fluctuations Toward Biological Functions, 2016Co-Authors: Shobhna Kapoor, Roland WinterAbstract:Fluctuations within biomolecules dictate a plethora of biological processes and are of great importance in functional studies in molecular biophysics. From shaping the free energy landscape of biomolecules themselves to that of biomolecular interactions, they are also implicated in a number of debilitating pathological diseases, thus generating exigent issues that require in-depth investigation. In this regard, Pressure Perturbation serves as an important tool to mechanistically explore the causes and effects of fluctuations in biomolecules and biomolecular assemblies. Here, we review the underlying principal action of Pressure on biomolecules with emphasis on lipid membranes, proteins, amyloids, and membrane-associated complexes along with some highlighted experiments. We first discuss how Pressure affects the structure, phase behavior, and dynamics of lipid membranes of varying complexity. We then review the promising role of this tool to study high-energy conformational and functional substates in proteins and elaborate on the use of Pressure modulation to understand protein aggregation and fibrillation phenomena. Finally, we present some recent results using Pressure Perturbation to explore membrane-associated biomolecular assemblies and uncover membrane-mediated conformational substates of proteins, furnishing unprecedented information on proteo-lipid interactions.
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Probing volumetric properties of biomolecular systems by Pressure Perturbation calorimetry (PPC)--the effects of hydration, cosolvents and crowding.
Methods (San Diego Calif.), 2014Co-Authors: Saba Suladze, Marie Kahse, Nelli Erwin, Daniel Tomazic, Roland WinterAbstract:Pressure Perturbation calorimetry (PPC) is an efficient technique to study the volumetric properties of biomolecules in solution. In PPC, the coefficient of thermal expansion of the partial volume of the biomolecule is deduced from the heat consumed or produced after small isothermal Pressure-jumps. The expansion coefficient strongly depends on the interaction of the biomolecule with the solvent or cosolvent as well as on its packing and internal dynamic properties. This technique, complemented with molecular acoustics and densimetry, provides valuable insights into the basic thermodynamic properties of solvation and volume effects accompanying interactions, reactions and phase transitions of biomolecular systems. After outlining the principles of the technique, we present representative examples on protein folding, including effects of cosolvents and crowding, together with a discussion of the interpretation, and further applications.
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Pressure Perturbation of actin suprastructures
Biophysical Journal, 2014Co-Authors: Christopher Rosin, Roland WinterAbstract:Parts of the cytoskeleton that are composed of actin are organized into filamentous cross-linked meshworks and bundles, which facilitate numerous cellular processes like cell migration, adhesion or cellular trafficking, to name a few. Actin bundles integrated into the cytoskeleton are key components for force generation and for reinforcing the cell against mechanical stress. From in vivo studies, the filamentous actin structures have been found to be among the most Pressure sensitive assemblies. Already in 1966, Ikkai and Ooi reported a strong sensitivity of cellular F-actin towards high hydrostatic Pressure (HHP), which depends on the presence of ATP and divalent cations. Details about the thermodynamics and structural changes of the Pressure-modulated G to F and F to G transition and HHP effects on actin suprastructures like networks and bundles are largely unknown, however. To shed more light on the stability profile of G-, F- and bundled actin, a variety of spectroscopic, microscopic and calorimetric methods was applied. The designed experiments reveal deeper insights into the structural and thermodynamic properties of these actin species over a wide range of temperatures and Pressures. A complete p,T-phase diagram, complemented by species-specific thermodynamic parameters, could be obtained, leading to a deeper molecular-level understanding of G-, filamentous and bundled actin, also at extreme environmental conditions.
George I. Makhatadze - One of the best experts on this subject based on the ideXlab platform.
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Applications of Pressure Perturbation calorimetry to study factors contributing to the volume changes upon protein unfolding.
Biochimica et biophysica acta, 2015Co-Authors: Pranav P. Pandharipande, George I. MakhatadzeAbstract:Abstract Background Pressure Perturbation calorimetry (PPC) is a biophysical method that allows direct determination of the volume changes upon conformational transitions in macromolecules. Scope of this review This review provides novel details of the use of PPC to analyze unfolding transitions in proteins. The emphasis is made on the data analysis as well as on the validation of different structural factors that define the volume changes upon unfolding. Four case studies are presented that show the application of these concepts to various protein systems. Major conclusions The major conclusions are: 1. Knowledge of the thermodynamic parameters for heat induced unfolding facilitates the analysis of the PPC profiles. 2. The changes in the thermal expansion coefficient upon unfolding appear to be temperature dependent. 3. Substitutions on the protein surface have negligible effects on the volume changes upon protein unfolding. 4. Structural plasticity of proteins defines the position dependent effect of amino acid substitutions of the residues buried in the native state. 5. Small proteins have positive volume changes upon unfolding which suggests difference in balance between the cavity/void volume in the native state and the hydration volume changes upon unfolding as compared to the large proteins that have negative volume changes. General significance The information provided here gives a better understanding and deeper insight into the role played by various factors in defining the volume changes upon protein unfolding. This article is part of a Special Issue entitled Microcalorimetry in the BioSciences — Principles and Applications, edited by Fadi Bou-Abdallah.
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Thermal expansivities of peptides, polypeptides and proteins as measured by Pressure Perturbation calorimetry.
Methods (San Diego Calif.), 2015Co-Authors: Pranav P. Pandharipande, George I. MakhatadzeAbstract:Abstract The main goal of this work was to provide direct experimental evidence that the expansivity of peptides, polypeptides and proteins as measured by Pressure Perturbation calorimetry (PPC), can serve as a proxy to characterize relative compactness of proteins, especially the denatured state ensemble. This is very important as currently only small angle X-ray scattering (SAXS), intrinsic viscosity and, to a lesser degree, fluorescence resonance transfer (FRET) experiments are capable of reporting on the compactness of denatured state ensembles. We combined the expansivity measurements with other biophysical methods (far-UV circular dichroism spectroscopy, differential scanning calorimetry, and small angle X-ray scattering). Three case studies of the effects of conformational changes on the expansivity of polypeptides in solution are presented. We have shown that expansivity appears to be insensitive to the helix–coil transition, and appears to reflect the changes in hydration of the side-chains. We also observed that the expansivity is sensitive to the global conformation of the polypeptide chain and thus can be potentially used to probe hydration of different collapsed states of denatured or even intrinsically disordered proteins.
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Molecular determinants of expansivity of native globular proteins: a Pressure Perturbation calorimetry study.
The journal of physical chemistry. B, 2014Co-Authors: Daniel Vasilchuk, Saba Suladze, Pranav P. Pandharipande, Jose M. Sanchez-ruiz, George I. MakhatadzeAbstract:There is a growing interest in understanding how hydrostatic Pressure (P) impacts the thermodynamic stability (ΔG) of globular proteins. The Pressure dependence of stability is defined by the change in volume upon denaturation, ΔV = (∂ΔG/∂P)T. The temperature dependence of change in volume upon denaturation itself is defined by the changes in thermal expansivity (ΔE), ΔE = (∂ΔV/∂T)P. The Pressure Perturbation calorimetry (PPC) allows direct experimental measurement of the thermal expansion coefficient, α = E/V, of a protein in the native, αN(T), and unfolded, αU(T), states as a function of temperature. We have shown previously that αU(T) is a nonlinear function of temperature but can be predicted well from the amino acid sequence using α(T) values for individual amino acids (J. Phys. Chem. B 2010, 114, 16166-16170). In this work, we report PPC results on a diverse set of nine proteins and discuss molecular factors that can potentially influence the thermal expansion coefficient, αN(T), and the thermal expansivity, EN(T), of proteins in the native state. Direct experimental measurements by PPC show that αN(T) and EN(T) functions vary significantly for different proteins. Using comparative analysis and site-directed mutagenesis, we have eliminated the role of various structural or thermodynamic properties of these proteins such as the number of amino acid residues, secondary structure content, packing density, electrostriction, dynamics, or thermostability. We have also shown that αN(T) and EN,sp(T) functions for a given protein are rather insensitive to the small changes in the amino acid sequence, suggesting that αN(T) and EN(T) functions might be defined by a topology of a given protein fold. This conclusion is supported by the similarity of αN(T) and EN(T) functions for six resurrected ancestral thioredoxins that vary in sequence but have very similar tertiary structure.
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Pressure Perturbation Calorimetry of Unfolded Proteins
The journal of physical chemistry. B, 2010Co-Authors: Alekos D. Tsamaloukas, Neena Pyzocha, George I. MakhatadzeAbstract:We report the application of Pressure Perturbation calorimetry (PPC) to study unfolded proteins. Using PPC we have measured the temperature dependence of the thermal expansion coefficient, α(T), in the unfolded state of apocytochrome C and reduced BPTI. We have shown that α(T) is a nonlinear function and decreases with increasing temperature. The decrease is most significant in the low (2−55 °C) temperature range. We have also tested an empirical additivity approach to predict α(T) of unfolded state from the amino acid sequence using α(T) values for individual amino acids. A comparison of the experimental and calculated functions shows a very good agreement, both in absolute values of α(T) and in its temperature dependence. Such an agreement suggests the applicability of using empirical calculations to predict α(T) of any unfolded protein.
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Use of Pressure Perturbation calorimetry to characterize the volumetric properties of proteins.
Methods in enzymology, 2009Co-Authors: Katrina L. Schweiker, George I. MakhatadzeAbstract:Pressure Perturbation calorimetry (PPC) is a new technique that makes possible to study the volumetric changes that occur upon protein unfolding. Here, we summarize the thermodynamic foundation of the method and introduce a two-state model for the analysis of the unfolding data monitored by PPC. Several examples of data analysis illustrating potential pitfalls and solutions are discussed.
Revanur Ravindra - One of the best experts on this subject based on the ideXlab platform.
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Pressure Perturbation calorimetric studies of the solvation properties and the thermal unfolding of proteins in solution experiments and theoretical interpretation
Physical Chemistry Chemical Physics, 2006Co-Authors: Lally Mitra, Catherine A. Royer, Revanur Ravindra, Nikolai Smolin, Roland WinterAbstract:We used Pressure Perturbation calorimetry (PPC), a relatively new and efficient technique, to study the solvation and volumetric properties of amino acids and peptides as well as of proteins in their native and unfolded state. In PPC, the coefficient of thermal expansion of the partial volume of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent or cosolvent at the protein–solvent interface. Furthermore, the effects of various chaotropic and kosmotropic cosolvents on the volume and expansivity changes of proteins were measured over a wide concentration range with high precision. Depending on the type of cosolvent and its concentration, specific differences were found for the solvation properties and unfolding behaviour of the proteins, and the volume change upon unfolding may even change sign. To yield a molecular interpretation of the different terms contributing to the partial protein volume and its temperature dependence, and hence a better understanding of the PPC data, molecular dynamics computer simulations on SNase were also carried out and compared with the experimental data. The PPC studies introduced aim to obtain more insight into the basic thermodynamic properties of protein solvation and volume effects accompanying structural transformations of proteins in various cosolvents on one hand, as these form the basis for understanding their physiological functions and their use in drug designing and formulations, but also to initiate further valuable applications in studies of other biomolecular and chemical systems.
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Pressure Perturbation calorimetric studies of the solvation properties and the thermal unfolding of proteins in solution—experiments and theoretical interpretation
Physical chemistry chemical physics : PCCP, 2006Co-Authors: Lally Mitra, Catherine A. Royer, Revanur Ravindra, Nikolai Smolin, Roland WinterAbstract:We used Pressure Perturbation calorimetry (PPC), a relatively new and efficient technique, to study the solvation and volumetric properties of amino acids and peptides as well as of proteins in their native and unfolded state. In PPC, the coefficient of thermal expansion of the partial volume of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent or cosolvent at the protein–solvent interface. Furthermore, the effects of various chaotropic and kosmotropic cosolvents on the volume and expansivity changes of proteins were measured over a wide concentration range with high precision. Depending on the type of cosolvent and its concentration, specific differences were found for the solvation properties and unfolding behaviour of the proteins, and the volume change upon unfolding may even change sign. To yield a molecular interpretation of the different terms contributing to the partial protein volume and its temperature dependence, and hence a better understanding of the PPC data, molecular dynamics computer simulations on SNase were also carried out and compared with the experimental data. The PPC studies introduced aim to obtain more insight into the basic thermodynamic properties of protein solvation and volume effects accompanying structural transformations of proteins in various cosolvents on one hand, as these form the basis for understanding their physiological functions and their use in drug designing and formulations, but also to initiate further valuable applications in studies of other biomolecular and chemical systems.
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Pressure Perturbation and differential scanning calorimetric studies of bipolar tetraether liposomes derived from the thermoacidophilic archaeon sulfolobus acidocaldarius
Biophysical Journal, 2005Co-Authors: Parkson Leegau Chong, Revanur Ravindra, Monika Khurana, Verrica English, Roland WinterAbstract:Differential scanning calorimetry (DSC) and Pressure Perturbation calorimetry (PPC) were used to characterize thermal phase transitions, membrane packing, and volumetric properties in multilamellar vesicles (MLVs) composed of the polar lipid fraction E (PLFE) isolated from the thermoacidophilic archaeon Sulfolobus acidocaldarius grown at different temperatures. For PLFE MLVs derived from cells grown at 78°C, the first DSC heating scan exhibits an endothermic transition at 46.7°C, a small hump near 60°C, and a broad exothermic transition at 78.5°C, whereas the PPC scan reveals two transitions at ∼45°C and 60°C. The endothermic peak at 46.7°C is attributed to a lamellar-to-lamellar phase transition and has an unusually low ΔH (3.5 kJ/mol) and ΔV/V (0.1%) value, as compared to those for the main phase transitions of saturated diacyl monopolar diester lipids. This result may arise from the restricted trans-gauche conformational changes in the dibiphytanyl chain due to the presence of cyclopentane rings and branched methyl groups and due to the spanning of the lipid molecules over the whole membrane. The exothermic peak at 78.5°C probably corresponds to a lamellar-to-cubic phase transition and exhibits a large and negative ΔH value (−23.2 kJ/mol), which is uncommon for normal lamellar-to-cubic phospholipid phase transformations. This exothermic transition disappears in the subsequent heating scans and thus may involve a metastable phase, which is irreversible at the scan rate used. Further, there is no distinct peak in the plot of the thermal expansion coefficient α versus temperature near 78.5°C, indicating that this lamellar-to-cubic phase transition is not accompanied by any significant volume change. For PLFE MLVs derived from cells grown at 65°C, similar DSC and PPC profiles and thermal history responses were obtained. However, the lower growth temperature yields a higher ΔV/V (∼0.25%) and ΔH (14 kJ/mol) value for the lamellar-to-lamellar phase transition measured at the same pH (2.1). A lower growth temperature also generates a less negative temperature dependence of α. The changes in ΔV/V, ΔH, and the temperature dependence of α can be attributed to the decrease in the number of cyclopentane rings in PLFE at the lower growth temperature. The relatively low ΔV/V and small ΔH involved in the phase transitions help to explain why PLFE liposomes are remarkably thermally stable and also echo the proposal that PLFE liposomes are generally rigid and tightly packed. These results help us to understand why, despite the occurrence of thermal-induced phase transitions, PLFE liposomes exhibit a remarkably low temperature sensitivity of proton permeation and dye leakage.
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hydration and packing effects on prion folding and β sheet conversion high Pressure spectroscopy and Pressure Perturbation calorimetry studies
Journal of Biological Chemistry, 2004Co-Authors: Yraima Cordeiro, Roland Winter, Revanur Ravindra, Debora Foguel, Julia Kraineva, Luis Mauricio T R Lima, Mariana P B Gomes, Jerson L. SilvaAbstract:Abstract The main hypothesis for prion diseases proposes that the cellular protein (PrPC) can be altered into a misfolded, β-sheet-rich isoform (PrPSc), which undergoes aggregation and triggers the onset of transmissible spongiform encephalopathies. Here, we compare the stability against Pressure and the thermomechanical properties of the α-helical and β-sheet conformations of recombinant murine prion protein, designated as α-rPrP and β-rPrP, respectively. High temperature induces aggregates and a large gain in intermolecular antiparallel β-sheet (β-rPrP), a conformation that shares structural similarity with PrPSc. α-rPrP is highly stable, and only Pressures above 5 kilobars (1 kilobar = 100 MegaPascals) cause reversible denaturation, a process that leads to a random and turnrich conformation with concomitant loss of α-helix, as measured by Fourier transform infrared spectroscopy. In contrast, aggregates of β-rPrP are very sensitive to Pressure, undergoing transition into a dissociated species that differs from the denatured form derived from α-rPrP. The higher susceptibility to Pressure of β-rPrP can be explained by its less hydrated structure. Pressure Perturbation calorimetry supports the view that the accessible surface area of α-rPrP is much higher than that of β-rPrP, which explains the lower degree of hydration of β-rPrP. Our findings shed new light on the mechanism of prion conversion and show how water plays a prominent role. Our results allow us to propose a volume and free energy diagram of the different species involved in the conversion and aggregation. The existence of different folded conformations as well as different denatured states of PrP may explain the elusive character of its conversion into a pathogenic form.
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Pressure Perturbation calorimetic studies of the solvation properties and the thermal unfolding of staphylococcal nuclease
Physical Chemistry Chemical Physics, 2004Co-Authors: Revanur Ravindra, Catherine A. Royer, Roland WinterAbstract:A rather new technique, Pressure Perturbation calorimetry (PPC), was applied to study volumetric and solvation properties of staphylococcal nuclease (Snase) in its native and unfolded state with high precision. Furthermore, the effects of various chaotropic and cosmotropic co-solvents on the solvation and unfolding behaviour of Snase was investigated in detail. In PPC, the apparent coefficient of thermal expansion of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent at the protein–solvent interface. In the native state, the protein shows a very strong thermal expansion of 1.0×10−3 K−1 at 10°C, which decreases steeply to 0.65×10−3 K−1 at 40°C. This behaviour is discussed in terms of a continuous release of condensed water from the protein surface. Upon unfolding, the volume decreases by about 19 mL×mol−1. Solutions of the cosmotropic and chaotropic compounds glycerol, sorbitol, K2SO4 and urea, respectively, show characteristic deviations from the thermal expansion and volumetric properties of the pure buffer solution. The solvent contribution to the apparent coefficient of thermal expansion of the protein, α, is enhanced considerably when the protein is immersed in a solvent known to be more structured than H2O (even the more structured D2O has a drastic effect) and nearly eliminated in a solvent in which “normal” water is largely absent (e.g., in 1.5 M urea). Similarly to D2O, a continuous increase in solvation was observed with increase in glycerol or sorbitol content in the buffer, which leads to an increase in protein stability, as is verified by the increasing Tm and ΔH values obtained by microcalorimetric measurements (DSC). In this regard, sorbitol is the more efficient agent. The reduction of ΔV in the presence of these stabilisers can in part be attributed to the formation of a partial unfolded state of the protein, in part it is due to the temperature dependence of ΔV. The preferential binding of urea reduces the hydration level, also in the native state, causing the protein to approach a more disordered state at high urea concentration. The increase in ΔV and the decrease in ΔH with increasing urea concentration support these conclusions. A stabilising effect, even though there is a reduction in solvation around the protein is observed for 0.5 M K2SO4 as co-solvent. In this case, surprisingly, ΔV is found to be positive which is an indication of the formation of a swollen, molten globule kind of unfolded state at the transition. Finally, ΔV values for the temperature-induced unfolding are compared with corresponding data for the Pressure-induced unfolding of Snase.
Catherine A. Royer - One of the best experts on this subject based on the ideXlab platform.
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Unique Features of the Folding Landscape of a Repeat Protein Revealed by Pressure Perturbation
Biophysical journal, 2010Co-Authors: Jean-baptiste Rouget, Roland Winter, Martin A. Schroer, Christoph Jeworrek, Matthias Pühse, Jean-louis Saldana, Yannick Bessin, Metin Tolan, Doug Barrick, Catherine A. RoyerAbstract:The volumetric properties of proteins yield information about the changes in packing and hydration between various states along the folding reaction coordinate and are also intimately linked to the energetics and dynamics of these conformations. These volumetric characteristics can be accessed via Pressure Perturbation methods. In this work, we report high-Pressure unfolding studies of the ankyrin domain of the Notch receptor (Nank1-7) using fluorescence, small-angle x-ray scattering, and Fourier transform infrared spectroscopy. Both equilibrium and Pressure-jump kinetic fluorescence experiments were consistent with a simple two-state folding/unfolding transition under Pressure, with a rather small volume change for unfolding compared to proteins of similar molecular weight. High-Pressure fluorescence, Fourier transform infrared spectroscopy, and small-angle x-ray scattering measurements revealed that increasing urea over a very small range leads to a more expanded Pressure unfolded state with a significant decrease in helical content. These observations underscore the conformational diversity of the unfolded-state basin. The temperature dependence of Pressure-jump fluorescence relaxation measurements demonstrated that at low temperatures, the folding transition state ensemble (TSE) lies close in volume to the folded state, consistent with significant dehydration at the barrier. In contrast, the thermal expansivity of the TSE was found to be equivalent to that of the unfolded state, indicating that the interactions that constrain the folded-state thermal expansivity have not been established at the folding barrier. This behavior reveals a high degree of plasticity of the TSE of Nank1-7.
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Pressure Perturbation calorimetric studies of the solvation properties and the thermal unfolding of proteins in solution experiments and theoretical interpretation
Physical Chemistry Chemical Physics, 2006Co-Authors: Lally Mitra, Catherine A. Royer, Revanur Ravindra, Nikolai Smolin, Roland WinterAbstract:We used Pressure Perturbation calorimetry (PPC), a relatively new and efficient technique, to study the solvation and volumetric properties of amino acids and peptides as well as of proteins in their native and unfolded state. In PPC, the coefficient of thermal expansion of the partial volume of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent or cosolvent at the protein–solvent interface. Furthermore, the effects of various chaotropic and kosmotropic cosolvents on the volume and expansivity changes of proteins were measured over a wide concentration range with high precision. Depending on the type of cosolvent and its concentration, specific differences were found for the solvation properties and unfolding behaviour of the proteins, and the volume change upon unfolding may even change sign. To yield a molecular interpretation of the different terms contributing to the partial protein volume and its temperature dependence, and hence a better understanding of the PPC data, molecular dynamics computer simulations on SNase were also carried out and compared with the experimental data. The PPC studies introduced aim to obtain more insight into the basic thermodynamic properties of protein solvation and volume effects accompanying structural transformations of proteins in various cosolvents on one hand, as these form the basis for understanding their physiological functions and their use in drug designing and formulations, but also to initiate further valuable applications in studies of other biomolecular and chemical systems.
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Pressure Perturbation calorimetric studies of the solvation properties and the thermal unfolding of proteins in solution—experiments and theoretical interpretation
Physical chemistry chemical physics : PCCP, 2006Co-Authors: Lally Mitra, Catherine A. Royer, Revanur Ravindra, Nikolai Smolin, Roland WinterAbstract:We used Pressure Perturbation calorimetry (PPC), a relatively new and efficient technique, to study the solvation and volumetric properties of amino acids and peptides as well as of proteins in their native and unfolded state. In PPC, the coefficient of thermal expansion of the partial volume of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent or cosolvent at the protein–solvent interface. Furthermore, the effects of various chaotropic and kosmotropic cosolvents on the volume and expansivity changes of proteins were measured over a wide concentration range with high precision. Depending on the type of cosolvent and its concentration, specific differences were found for the solvation properties and unfolding behaviour of the proteins, and the volume change upon unfolding may even change sign. To yield a molecular interpretation of the different terms contributing to the partial protein volume and its temperature dependence, and hence a better understanding of the PPC data, molecular dynamics computer simulations on SNase were also carried out and compared with the experimental data. The PPC studies introduced aim to obtain more insight into the basic thermodynamic properties of protein solvation and volume effects accompanying structural transformations of proteins in various cosolvents on one hand, as these form the basis for understanding their physiological functions and their use in drug designing and formulations, but also to initiate further valuable applications in studies of other biomolecular and chemical systems.
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Pressure Perturbation calorimetic studies of the solvation properties and the thermal unfolding of staphylococcal nuclease
Physical Chemistry Chemical Physics, 2004Co-Authors: Revanur Ravindra, Catherine A. Royer, Roland WinterAbstract:A rather new technique, Pressure Perturbation calorimetry (PPC), was applied to study volumetric and solvation properties of staphylococcal nuclease (Snase) in its native and unfolded state with high precision. Furthermore, the effects of various chaotropic and cosmotropic co-solvents on the solvation and unfolding behaviour of Snase was investigated in detail. In PPC, the apparent coefficient of thermal expansion of the protein is deduced from the heat consumed or produced after small isothermal Pressure jumps, which strongly depends on the interaction of the protein with the solvent at the protein–solvent interface. In the native state, the protein shows a very strong thermal expansion of 1.0×10−3 K−1 at 10°C, which decreases steeply to 0.65×10−3 K−1 at 40°C. This behaviour is discussed in terms of a continuous release of condensed water from the protein surface. Upon unfolding, the volume decreases by about 19 mL×mol−1. Solutions of the cosmotropic and chaotropic compounds glycerol, sorbitol, K2SO4 and urea, respectively, show characteristic deviations from the thermal expansion and volumetric properties of the pure buffer solution. The solvent contribution to the apparent coefficient of thermal expansion of the protein, α, is enhanced considerably when the protein is immersed in a solvent known to be more structured than H2O (even the more structured D2O has a drastic effect) and nearly eliminated in a solvent in which “normal” water is largely absent (e.g., in 1.5 M urea). Similarly to D2O, a continuous increase in solvation was observed with increase in glycerol or sorbitol content in the buffer, which leads to an increase in protein stability, as is verified by the increasing Tm and ΔH values obtained by microcalorimetric measurements (DSC). In this regard, sorbitol is the more efficient agent. The reduction of ΔV in the presence of these stabilisers can in part be attributed to the formation of a partial unfolded state of the protein, in part it is due to the temperature dependence of ΔV. The preferential binding of urea reduces the hydration level, also in the native state, causing the protein to approach a more disordered state at high urea concentration. The increase in ΔV and the decrease in ΔH with increasing urea concentration support these conclusions. A stabilising effect, even though there is a reduction in solvation around the protein is observed for 0.5 M K2SO4 as co-solvent. In this case, surprisingly, ΔV is found to be positive which is an indication of the formation of a swollen, molten globule kind of unfolded state at the transition. Finally, ΔV values for the temperature-induced unfolding are compared with corresponding data for the Pressure-induced unfolding of Snase.
Robert J. Falconer - One of the best experts on this subject based on the ideXlab platform.
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analysis of mesoscopic structured 2 propanol water mixtures using Pressure Perturbation calorimetry and molecular dynamic simulation
Journal of Solution Chemistry, 2017Co-Authors: Jordan W. Bye, Colin L. Freeman, John D. Howard, Gregor Herz, James Mcgregor, Robert J. FalconerAbstract:In this paper we demonstrate the application of Pressure Perturbation calorimetry (PPC) to the characterization of 2-propanol/water mixtures. PPC of different 2-propanol/water mixtures provides two useful measurements: (i) the change in heat (ΔQ); and (ii) the \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) value. The results demonstrate that the ΔQ values of the mixtures deviate from that expected for a random mixture, with a maximum at ~20–25 mol% 2-propanol. This coincides with the concentration at which molecular dynamics (MD) simulations show a maximum deviation from random distribution, and also the point at which alcohol–alcohol hydrogen bonds become dominant over alcohol–water hydrogen bonds. Furthermore, the \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) value showed transitions at 2.5 mol% 2-propanol and at approximately 14 mol% 2-propanol. Below 2.5 mol% 2-propanol the values of \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) are negative; this is indicative of the presence of isolated 2-propanol molecules surrounded by water molecules. Above 2.5 mol% 2-propanol \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) rises, reaching a maximum at ~14 mol% corresponding to a point where mixed alcohol–water networks are thought to dominate. The values and trends identified by PPC show excellent agreement not only with those obtained from MD simulations but also with results in the literature derived using viscometry, THz spectroscopy, NMR and neutron diffraction.
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Analysis of Mesoscopic Structured 2-Propanol/Water Mixtures Using Pressure Perturbation Calorimetry and Molecular Dynamic Simulation
Journal of solution chemistry, 2016Co-Authors: Jordan W. Bye, Colin L. Freeman, John D. Howard, Gregor Herz, James Mcgregor, Robert J. FalconerAbstract:In this paper we demonstrate the application of Pressure Perturbation calorimetry (PPC) to the characterization of 2-propanol/water mixtures. PPC of different 2-propanol/water mixtures provides two useful measurements: (i) the change in heat (ΔQ); and (ii) the \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) value. The results demonstrate that the ΔQ values of the mixtures deviate from that expected for a random mixture, with a maximum at ~20–25 mol% 2-propanol. This coincides with the concentration at which molecular dynamics (MD) simulations show a maximum deviation from random distribution, and also the point at which alcohol–alcohol hydrogen bonds become dominant over alcohol–water hydrogen bonds. Furthermore, the \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) value showed transitions at 2.5 mol% 2-propanol and at approximately 14 mol% 2-propanol. Below 2.5 mol% 2-propanol the values of \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) are negative; this is indicative of the presence of isolated 2-propanol molecules surrounded by water molecules. Above 2.5 mol% 2-propanol \( \left[ {\delta \bar{C}_{p} /\delta p} \right]_{T} \) rises, reaching a maximum at ~14 mol% corresponding to a point where mixed alcohol–water networks are thought to dominate. The values and trends identified by PPC show excellent agreement not only with those obtained from MD simulations but also with results in the literature derived using viscometry, THz spectroscopy, NMR and neutron diffraction.
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A study of the relationship between water and anions of the Hofmeister series using Pressure Perturbation calorimetry
Physical chemistry chemical physics : PCCP, 2015Co-Authors: Jordan W. Bye, Robert J. FalconerAbstract:Pressure Perturbation calorimetry (PPC) was used to study the relationship between water and sodium salts with a range of different anions. At temperatures around 25 °C the heat on pressurisation (ΔQ) from 1 to 5 bar was negative for all solutions relative to pure water. The raw data showed that as the temperature rose, the gradient was positive relative to pure water and the transition temperature where ΔQ was zero was related to anion surface charge density and was more pronounced for the low-charge density anions. A three component model was developed comprising bulk water, the hydration layer and the solute to calculate the molar expansivity of the hydration layer around the ions in solution. The calculated molar expansivities of water in the hydration layer around the ions were consistently less than pure water. ΔQ at different disodium hydrogen phosphate concentrations showed that the change in molar enthalpy relative to pure water was not linear even as it approached infinite dilution suggesting that while hydration layers can be allocated to the water around ions this does not rule out interactions between water and ions extending beyond the immediate hydration layer.