Preferential Interaction Parameter Research Articles

Preferential Solvation Phenomena as Solute-Induced Structure-Making/Breaking Processes: Linking Thermodynamic Preferential Interaction Parameters to Fundamental Structure Making/Breaking Functions.

In this work, we identify the explicit macroscopic-to-microscopic rigorous links between existing thermodynamic preferential interaction parameters and microstructural descriptors based on total correlation function integrals, leading to their unambiguous characterization in terms of fundamental structure making/breaking functions . First, we provide the statistics mechanical framework to identify a universal molecular-based signature for the preferential solvation phenomenon involving solutes at infinite dilution in mixed-solvent environments and discuss its fundamental properties. Then, we characterize the functions relevant to the process, identify the microscopic markers for the existing preferential interaction parameters in terms of the functions, and test their compliance with a pair of essential microstructural constraints linked to the properties of the universal signature. Moreover, we illustrate the analysis by probing the behavior of a representative ternary system comprising the solubility of methane in aqueous 1,4-dioxane mixed-solvent environments under ambient conditions. Finally, we discuss some relevant issues surrounding the statistical mechanical (microstructural) interpretation of the thermodynamic (macroscopic) preferential interaction parameters, review some pitfalls in their evaluation from molecular simulation, and provide an outlook.

Trifluoroethanol direct interactions with protein backbones destabilize α-helices

2,2,2-Trifluoroethanol (TFE) is a well-known protein α-helix stabilizer; nevertheless, after much investigation, no consensus has been reached on TFE's stabilizing mechanism. TFE alters the structure of water and affects its dielectric properties, but also competes with it for hydrogen bonds with the backbone and side chains of proteins. Thus, indirect and direct mechanisms of TFE activity have been proposed. The direct mode is especially appealing: TFE establishes hydrogen bonds with the carbonyls of the peptides' backbones, eliminating water, apparently protecting the intra-helix hydrogen bond. Because these interactions occur simultaneously with other changes in the solution structure, it is difficult to disentangle the contribution of direct vs. indirect processes to the TFE stabilizing effect. Here, we perform extensive enhanced sampling simulations of the (AAQAA)3 peptide in mixtures of water and TFE at various concentrations. Minimum-distance distribution functions (MDDFs) and the Kirkwood-Buff (KB) theory of solutions are used to understand the molecular and thermodynamic basis of the TFE mechanism of α-helix stabilization. The simulations confirm the stabilizing role of TFE on the helical content of the peptide and that the helical structures are preferentially solvated by TFE. TFE effectively interacts with the protein backbone, excluding water, in agreement with the direct-interaction model. Yet, simulations allow alchemical experiments to be performed, and thus we modified the intermolecular backbone-TFE interactions to prevent the putatively stabilizing hydrogen-bond. Surprisingly, the peptide's helical content increased, showing that these direct contacts have a denaturing effective contribution. At the same time, the preferential interaction parameters remain basically constant in the absence of the TFE-backbone hydrogen bonds. Therefore, the model of TFE helix stabilization based on the protection of backbone hydrogen bonds by hydrogen-bonding the backbone’s carbonyl is not supported by evidence. We show that TFE non-specific interactions with the helical conformations are stronger than with the coil states, excluding water.

An able-cryoprotectant and a moderate denaturant: distinctive character of ethylene glycol on protein stability

Osmolytes are known to stabilize proteins against denaturing conditions. Ethylene glycol (EG), however, shows a distinctive effect on α-lactalbumin (α-LA) that it stabilizes the protein against cold-induced denaturation, whereas it destabilizes during heat denaturation. The replica exchange molecular dynamics (REMD) simulation of α-LA in the presence of EG shows that EG denatures the protein at higher temperatures whereas it retards the denaturation at sub-zero temperature. Representative structures of α-LA were selected from REMD trajectories at three different temperature conditions (240, 300 and 340 K) with and without EG, and classical molecular dynamics (MD) simulations were performed. The results suggest that the presence of water around α-LA is more at lower temperatures; however, water around the hydrophobic residues is reduced with the addition of EG at sub-zero temperature. The partition coefficient of EG showed that the binding of EG with hydrophobic residues was higher at lower temperatures. Preferential interaction parameters at different temperatures were calculated based on the mean distribution (Γ23) and Kirkwood–Buff integral (G 23) methods. Γ23 shows a larger positive value at 240 K compared to higher temperatures. G 23 shows positive values at lower temperatures, whereas it becomes negative at above 280 K. These results indicate that the preferential binding of EG with α-LA is more at sub-zero temperature compared to higher temperature conditions. Thus, the study suggests that the preferential binding of EG reduces the hydrophobic hydration of α-LA at lower temperatures, and stabilizes the protein against cold denaturation. However, the preferential binding of EG at higher temperature drives the folding equilibrium towards the denatured state. Communicated by Ramaswamy H. Sarma

Investigation on the Mechanisms of Synchronous Interaction of K3Cit with Melamine and Uric Acid That Avoids the Formation of Large Clusters.

Uric acid (UA) has an enormous competence to aggregate over melamine (Mel), producing large UA clusters that "drag" Mel toward them. Such a combination of donor-acceptor pairs provides a robust Mel-UA composite, thereby denoting a high complexity. Thus, a straightforward but pragmatic methodology might indeed require either destruction of the aggregation of UA or impediment of the hydrogen-bonded cluster of Mel and UA. Here, potassium citrate (K3Cit) is used as a potent inhibitor for a significant decrease of large UA-Mel clusters. The underlying mechanisms of synchronous interactions between K3Cit and the Mel-UA pair are examined by the classical molecular dynamics simulation coupled with the enhanced sampling method. K3Cit binds to the Mel-UA pair profoundly to produce a Mel-UA-K3Cit complex with favorable complexation energy (as indicated by the reckoning of pairwise ΔGbind° employing the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method). The strength of interaction follows the order UA-K3Cit > Mel-K3Cit > Mel-UA, thus clearly demonstrating the instability caused by upsetting the π-stacking of UA and hydrogen bonding of Mel-UA simultaneously. The comprehensive, strategically designed "direct approach" and "indirect approach" cluster structure analysis shows that K3Cit reduces the direct approach Mel-UA cluster size significantly irrespective of ensemble variation. Furthermore, the estimation of potentials of mean force (PMFs) reveals that the (UA)decamer-Mel interaction prevails over (UA)tetramer-Mel. The dynamic property (dimer existence autocorrelation functions) proves the essence of dimerization between Mel and UA in the absence and presence of K3Cit. Moreover, the calculation of the preferential interaction parameter provides the concentration at which Mel-K3Cit and UA-K3Cit interactions are predominant over the interaction of Mel and UA.

Preferential Interaction of Chymotrypsinogen in Aqueous Solutions of Polyols at 298.15 K

Preferential interactions of α-chymotrypsinogen in water and aqueous solutions of ethylene glycol, glycerol, erythritol, sorbitol, and inositol were studied from precise density measurements under conditions of constant molality and constant chemical potential; to achieve the last condition, solutions were brought under equilibrium dialysis. The density measurements were performed at 298.15 K and they were used to determine the preferential interaction parameters. The results confirm a significant correlation between the numbers of hydroxyl groups of the polyols with the stability of α-chymotrypsinogen.

Inclusion of Theobromine Modifies Uric Acid Aggregation with Possible Changes in Melamine–Uric Acid Clusters Responsible for Kidney Stones

Theobromine, a naturally occurring substance, can be conceived as a prospective inhibitor for uric acid clustering. In aqueous solution, aggregates of π-stacked uric acid molecules with the larger size of clusters are modified into lower-order clusters with a substantial percentage of monomer by the incorporation of theobromine. The composite made of theobromine-uric acid is expected to have enhanced water solubility, allowing stable kidney stones to be excreted through urine. Interestingly, the strategy for the decomposition with feasible modifications in melamine-uric acid composites (that are hydrogen-bonded) is developed (by implementing the cluster structure analysis technique and binding free energies). The all-atom molecular dynamics (MD) data provides new insights into the structure and dynamics of uric acid along with melamine molecules in the context of aggregation. The simulation in the present study is supported further by structural and dynamical property calculations. The calculations of hydrogen bond dynamics, the average number of hydrogen bonds, dimer existence autocorrelation functions, umbrella sampling, and coordination number theorize that the incorporation of theobromine significantly modifies the aggregated structure of uric acid. The overall complexation energy, along with the quantum chemical calculations, further explains the alternation of aggregated structure. Furthermore, the preferential interaction parameter describes at which concentration theobromine-uric acid interaction (which is π-stacked) predominates over uric acid-uric acid interactions. Interestingly, the interactions between theobromine-melamine and melamine-melamine (which are hydrogen-bonded) are not relevant here. Thus, melamine-uric acid cluster size is reduced owing to the disintegration of self-aggregated uric acid clusters by the involvement of theobromine. Moreover, an excellent agreement is observed between present MD results and experimentally obtained data.

Theoretical Insight into Preferential Interaction Issues and Solution Structure, and Contentious Apparent Hydrated Molar Volume of Cosolute

Background: There seems to be a mathematical or a conceptual error in an equation whose substitution into other equations for the determination of an apparent hydrated molar volume (V1) of a cosolute leads to an incorrect answer.
 Objectives: The objectives are 1) To show theoretically that the preferential interaction parameter (PIP) is an extensive thermodynamic quantity, 2) rederive new equations and reexamine various equations related to solution structure, 3) apply derived equation for the determination of V1, and 4) determine m-values and cognate preferential interaction parameter (PIP).
 Methods: The research is mainly theoretical and partly experimental. Bernfeld method of enzyme assay was adopted for the generation of data.
 Results and Discussion: The investigation showed that equation linking chemical potential of osmolyte to solution structure is dimensionally invalid; PIP was seen as a thermodynamically extensive quantity. Equations for the graphical determination of V1 of the osmolyte were derived. With ethanol alone, there were - m-value and + PIP; with aspirin alone, there were + m-value and - PIP. There was a change in sign in m-value with sucrose and ethanol/aspirin mixture, and a change in sign in PIP when the latter is taken as function of [ethanol]/[aspirin] and [sucrose](c3).
 Conclusion: A solution structure is as usual determined by either a relative excess or a deficit of the solution component either in the bulk or around the macromolecular surface domain; the PIP remains thermodynamically an extensive quantity. To be valid there is a need to introduce a reference standard molar concentration or activity to some equations in literature. The slope from one of the equations seems to give a valid value for V1 (V1 is «1; is activity coefficient). A known destabiliser may behave as a stabiliser being excluded. Like ethanol, aspirin as cosolute is destabilising and opposed by sucrose.

Underlying mechanistic insights into the structural properties of melamine and uric acid complexes with compositional variation under ambient conditions

The structural properties of melamine-uric acid complexes (which are responsible for kidney stones) with compositional variations are examined using a series of classical molecular dynamics simulations. The preferential interaction parameters imply that melamine interacts more strongly with uric acid than with other melamine molecules present in the system, whereas uric acid preferentially interacts with other uric acid molecules rather than with melamine. The stronger interactions among uric acid molecules produce higher-order uric acid clusters, which “drag” neighboring melamine molecules to be added to a cluster. Determination of orientational preferences between aromatic planes reveals that π–π stacking is responsible for uric acid self-association but less significant for melamine-melamine and melamine-uric acid accumulation. Cluster structure analyses suggest that higher concentrations of melamine, uric acid, or both result in a large insoluble melamine-uric acid complex cluster. Molecular mechanics-Poisson Boltzmann surface area calculations give a negative binding energy, indicating favorable complexation between melamine and uric acid molecules. Moreover, the overall complexation energy [ΔG0(mel-mel)+ ΔG0(uri-uri)+ ΔG0(mel-uri)] is more negative than ΔG0bind(mel-uri). The lifetime of melamine dimers is quite low compared with those of uric acid-uric acid and melamine-uric acid dimers, resulting in a low percentage of larger clusters for melamine-melamine interaction and a significant percentage of higher-order melamine-uric acid and uric acid-uric acid clusters with longer lifetimes. Furthermore, melamine and uric acid form strong hydrogen bonds, and melamine-melamine interactions are dominated by hydrogen bonding, whereas uric acid forms only a small number of hydrogen bonds with other uric acid molecules.

Activity Coefficient of Solution Components and Salts as Special Osmolyte from Kirkwood-Buff Theoretical Perspective

Background: There has been different interpretation of kosmotropes and chaotropes without concern for the physicochemical characteristics of the macromolecule and for the link between Hofmeister phenomena with solution structure. The objectives of this research are: 1) To investigate different ways of determining activity coefficient and activity of ionic osmolyte 2), to present a common theoretical basis for the interaction between reaction mixture components and Hofmeister phenomena and 3) determine the preferential interaction parameters and the Kirkwood-Buff integrals.
 Methods: A major theoretical research and partly experimental.
 Results and Discussion: Some equations in literature gave different values of activity coefficient and activity of solution components. The preferential interaction by binding is positive with ethanol only and at its higher concentration in the presence of ideal solution of different concentration of calcium chloride. There was positive m-value with ethanol. It was negative m-value in the presence of preferentially binding species, calcium ion and ethanol as against the excluded chloride ion. There was negative and positive change of solvation preference and interaction parameter due respectively to ethanol only and a mixture of it and the salt.
 Conclusion: Selected equations in literature may not give the same values of activity coefficient and activity of solution components. The presence of stabilising osmolyte, salt, and ethanol may not always yield positive m-values. The sign of the change of solvation preference with either binary or ternary mixture of osmolytes and, the cognate interaction parameter, may be a better indicator of the stability of a macromolecule. The kosmotropes and chaotropes may be cationic or anionic and their deficit or otherwise around the macromolecule and consequence, depend largely on net charge on the macromolecule at a given pH.

How does the complexation ability between host endo-functionalized molecular tube and strongly hydrophilic guest molecules in water depend on guest concentration?

We have already shown in our previous work (R. Paul, S. Paul, Synergistic Host-Guest Hydrophobic and Hydrogen Bonding Interactions in the Complexation between Endo-Functionalized Molecular Tube and Strongly Hydrophilic Guest Molecules in Aqueous Solution, Phys. Chem. Chem. Phys. 20 (2018) 16540–16550.) the main factors responsible in the complexation between an endo-functionalized molecular tube and different strongly hydrophilic guest molecules in water. These types of molecular tubes have gained a significant importance for encapsulating many environmentally and biologically important molecules. Here, we have examined how does this type of complexation ability depend on concentration of guest molecules? We use classical molecular dynamics together with MM-PBSA method for this purpose. We examine the effect of concentration of different guest molecules on the host-guest complexation ability by determining different types of hydrogen bond properties, preferential interaction parameter, water-inserted guest interaction energy parameter, host-guest binding free energy and potential of mean force. From the calculated free energy values obtained from MM-PBSA and PMF calculation, it is revealed that as the concentration of guest decreases the free energy becomes less favorable.

Basic Kirkwood – Buff Theory of Solution Structure and Appropriate Application of Wyman Linkage Equation to Biochemical Phenomena

Background: Researchers who have shown interest in the consequence of introducing dry biomolecules or a solution of it into cosolvents generally known as osmolyte, have applied many models for the elucidation of the scientific basis of the results obtained. The Kirkwood and Buff theory (KBT) or its reverse form has been the basis for the interpretation of the effect of the osmolyte. There seems to be no generally acceptable definition of terms in the basic KBT mathematical formalism. There is also error in stated equations describing solution structure and misapplication of Wyman linkage relation. Therefore, the objectives of this research are 1) to show how the equation of preferential interaction parameter is derived based on KBT, 2) to show the appropriate way in which Wyman linkage relation can be applied, 3) to apply biochemical approach (using generated data) to the equation of preferential interaction parameter (preferential interaction parameter is symbolised as ) for its calculation and calculation of parameters linked to KBT derived equations.
 Methods: The research is mainly theoretical and partly experimental. The experiment entails Bernfeld method of enzyme assay for the generation of data.
 Results and Discussion: The change in solvation preference upon the ethanol partial denaturation of the enzyme and the corresponding change in preferential interaction parameter were respectively positive and negative in sign. Unexpectedly ethanol was preferentially excluded from the enzyme.
 Conclusion: The equations of preferential interaction parameters were derived. The appropriate way is either by calculation or measurement of preferential interaction parameter. Therefore, or for the change, cannot be a constant (or slope) and an instrumentation–based measurable parameter at the same time. Based on Wyman linkage relation, purely biochemical thermodynamic parameter is linked to preferential interaction parameters which are therefore, thermodynamic parameters.

How does temperature modulate the structural properties of aggregated melamine in aqueous solution-An answer from classical molecular dynamics simulation.

In this study, classical molecular dynamics simulation of eight melamine molecules is carried out in water over a temperature range of 300 K to 380 K at an ambient pressure to examine the molecular details of melamine aggregation along with the impact of temperature on the aggregated state of melamine in water. It is found that the hydrogen bonds formed between sp3 N-sp2 N of melamine, which is mainly responsible for the aggregation over the sp3 N-sp3 N, are disturbed mainly by the rise in temperature. These outcomes are complemented by the consideration of an average number of hydrogen bonds per melamine and preferential interaction parameter calculations. The impact of temperature is negligible on the orientational probability between the two triazine cores. The π-π stacking interaction between the two triazine rings plays a less significant role on melamine aggregation. Dynamical calculations, by considering cluster structure analyses and dimer existence autocorrelation function, strengthen the fact of destabilization of aggregated melamine in water with the rise in temperature. With free energy of solvation, association constant along with the binding free energy between a melamine pair gives the thermodynamic point of view of the impact of elevated temperature on melamine aggregation. Interestingly, the potential of mean force calculation using an umbrella sampling technique explains the reasons, in depth, of how do sp3 N-sp2 N interactions confirm the decrease in the initial probability of growth of higher order clusters with the increase in temperature.

Preferential Solvation/Hydration of α-Chymotrypsin in Water-Acetonitrile Mixtures.

The aim of our study is to monitor the preferential hydration/solvation of the protein macromolecules at low and high water content in water-organic mixtures. Our approach is based on the analysis of the absolute values of the water/organic solvent sorption. We applied this approach to estimate the protein stabilization/destabilization due to the preferential interactions of α-chymotrypsin with water-acetonitrile mixtures. At high water content, α-chymotrypsin is preferentially hydrated. At the intermediate water content, the preferential interaction changed from preferential hydration to preferential binding of acetonitrile. From infrared spectra, changes in the structure of α-chymotrypsin were determined through an analysis of the structure of the amide I band. Acetonitrile augments the intensity of the 1626 cm-1 band assigned to the intermolecular β-sheet aggregates. At low water content, the protein is in a glassy (rigid) state. The H-bond accepting acetonitrile molecules are not effective in solvating the dehydrated protein molecules alone. Therefore, the acetonitrile molecules are preferentially excluded from the protein surface, resulting in the preferential hydration. Advantages of our approach: (i) The preferential interaction parameters can be determined in the entire range of water content in water-organic mixtures. (ii) Our approach facilitates the individual evaluation of the Gibbs energies of water, protein, and organic solvent.

Role of caffeine as an inhibitor in aggregation of hydrophobic molecules: A molecular dynamics simulation study

Hydrophobic effect is one of the major driving forces in biomolecular interactions. Biological molecules such as proteins and nucleic acids contain numerous hydrophobic components, and their interactions play critical roles in protein folding-unfolding and protein aggregation. In our study, we have carried out molecular dynamics simulation to understand the effect of caffeine on hydrophobic aggregation of di-t-butyl-methane (DTBM) molecules. For this we consider aqueous solutions of a regime of caffeine:DTBM stoichiometric ratios. From the calculations of site-site radial distribution functions followed by the coordination number analyses and spatial distribution plots we have observed the disruption of hydrophobic moieties of DTBM aggregates in 10:1 or more stoichiometric ratio of caffeine:DTBM systems. Calculations of binding affinities, preferential interaction parameters, and effect of caffeine cluster's size on aggregation show that though the binding affinity of DTBM with itself is more when compared to that of caffeine-DTBM, with increasing caffeine number the former decreases. On the other hand, in the presence of higher number of caffeine molecules the formation of caffeine clusters takes place and these caffeine clusters form a hydrophobic environment in which DTBM molecule is encapsulated. The presence of a significant number of water molecules in this confinement also ensures the hydration of DTBM. Moreover, these caffeine clusters also act as barriers and physically block the DTBM molecules to interact with the like molecules leading to a disruption of DTBM aggregates in caffeine solution.

Competitive interaction of monovalent cations with DNA from 3D-RISM.

The composition of the ion atmosphere surrounding nucleic acids affects their folding, condensation and binding to other molecules. It is thus of fundamental importance to gain predictive insight into the formation of the ion atmosphere and thermodynamic consequences when varying ionic conditions. An early step toward this goal is to benchmark computational models against quantitative experimental measurements. Herein, we test the ability of the three dimensional reference interaction site model (3D-RISM) to reproduce preferential interaction parameters determined from ion counting (IC) experiments for mixed alkali chlorides and dsDNA. Calculations agree well with experiment with slight deviations for salt concentrations >200 mM and capture the observed trend where the extent of cation accumulation around the DNA varies inversely with its ionic size. Ion distributions indicate that the smaller, more competitive cations accumulate to a greater extent near the phosphoryl groups, penetrating deeper into the grooves. In accord with experiment, calculated IC profiles do not vary with sequence, although the predicted ion distributions in the grooves are sequence and ion size dependent. Calculations on other nucleic acid conformations predict that the variation in linear charge density has a minor effect on the extent of cation competition.

Exploring the Counteracting Mechanism of Trehalose on Urea Conferred Protein Denaturation: A Molecular Dynamics Simulation Study.

To provide the underlying mechanism of the inhibiting effect of trehalose on the urea denatured protein, we perform classical molecular dynamics simulations of N-methylacetamide (NMA) in aqueous urea and/or trehalose solution. The site-site radial distribution functions and hydrogen bond properties indicate in binary urea solution the replacement of NMA-water hydrogen bonds by NMA-urea hydrogen bonds. On the other hand, in ternary urea and trehalose solution, trehalose does not replace the NMA-urea hydrogen bonds significantly; rather, it forms hydrogen bonds with the NMA molecule. The calculation of a preferential interaction parameter shows that, at the NMA surface, trehalose molecules are preferred and the preference for urea decreases slightly in ternary solution with respect to the binary solution. The exclusion of urea molecules in the ternary urea-NMA-trehalose system causes alleviation in van der Waals interaction energy between urea and NMA molecules. Our findings also reveal the following: (a) trehalose and urea induced second shell collapse of water structure, (b) a reduction in the mean trehalose cluster size in ternary solution, and (c) slowing down of translational motion of solution species in the presence of osmolytes. Implications of these results for the molecular explanations of the counteracting mechanism of trehalose on urea induced protein denaturation are discussed.

Understanding the role of temperature change and the presence of NaCl salts on caffeine aggregation in aqueous solution: from structural and thermodynamics point of view.

To examine the molecular level understanding of temperature induced self-association of caffeine molecules in aqueous solution both in the presence and absence of salt NaCl, we have performed long MD simulations at a regime of temperatures ranging from 275 to 350 K with a temperature difference of 25 K. The calculations of different site-site radial distribution functions followed by coordination number analyses, the calculations of preferential interaction parameters, solvent accessible surface area, and cluster structure analyses show a depletion in the caffeine association propensity with increasing temperature. We have also observed the salting out effect of caffeine molecules in salt solution. The simultaneous presence of polar and nonpolar groups in a caffeine molecule leads to anisotropic hydration. Specifically, the hydration tendency of caffeine hydrophobic sites increases with increasing temperature, while hydrophilic sites tend to be less hydrated. This leads to a decrease in caffeine association. In accordance with some experimental studies on thermodynamics of caffeine association, we have also observed enthalpy driven association in pure water. But the presence of salt leads to entropy driven association specifically at higher temperature. This is due to the relatively stronger interactions of salt ions with caffeine at higher temperature.

Molecular insights into the role of aqueous trehalose solution on temperature-induced protein denaturation.

To investigate the underlying mechanism by which trehalose acts as a bioprotectant against thermal denaturation of protein in aqueous solution, we carry out classical molecular dynamics simulations at two different temperatures. Though it is widely accepted that trehalose acts as an antidote against such protein structural destabilization and numerous hypotheses have been proposed in regard to its mechanism of stabilization, there is still no definitive generally accepted answer to this question and it remains a subject of active research. In view of this, in this article we report the thermal denaturation process of a 15-residue S-peptide analogue at 360 K temperature and the counteracting ability of trehalose of varying concentrations at that temperature. In order to verify the conformational stability of the peptide at ambient temperature condition, we also carry out a separate simulation of peptide-water binary system at 300 K temperature. The goal is to provide a molecular level understanding of how trehalose protects protein at elevated temperature. The Cα-rmsd calculation shows that in pure water, the peptide is stable at 300 K temperature and its unfolding is observed at 360 K. However, in peptide-water-trehalose ternary system, the value of Cα-rmsd decreases as trehalose concentration is increased. Remarkably, at the highest trehalose concentration considered in this study, the value of Cα-rmsd at 360 K is similar to that of water-peptide binary system at 300 K temperature. Further, the calculations of radius of gyration of Cα-atoms and helical percentage of the peptide residues support the above observations. The total number of hydrogen bonds formed by the peptide with solution species (trehalose and water) remains constant, though the peptide water hydrogen bond decreases and peptide trehalose hydrogen bond increases with increasing trehalose concentration. This finding suggests replacement of water molecules by trehalose molecules and supports water replacement hypothesis. The calculations of preferential interaction parameter show that at the peptide surface, trehalose molecules are slightly more preferred over water and for the most concentrated solutions, a prominent exclusion of water and enrichment of trehalose molecules is observed. Also observed are (i) trehalose-induced second shell collapse of water structure, (ii) the growth of trehalose cluster as concentration is increased, and (iii) trehalose-induced slowing down of the translational motion of both water and trehalose, the effect being more pronounced for the latter. Implications of these results for counteracting mechanism of trehalose are discussed.

Revisiting the conundrum of trehalose stabilization.

Protein aggregation and loss of protein's biological functionality are manifestations of protein instability. Cosolvents, in particular trehalose, are widely accepted antidotes against such destabilization. Although numerous theories have been promulgated in the literature with regard to its mechanism of stabilization, the present scenario is still elusive in view of the discrepancies existing in them. To this end, we have revisited the conundrum and attempted to rationalize the mechanism by conducting thorough investigation of the effect of trehalose on the native, partially unfolded and denatured states of protein "Lysozyme" by means of molecular dynamic (MD) simulations under different temperature and concentration regimes. Two-dimensional contour plots along with principal component analysis suggest that trehalose molecules offer on-pathway stabilization unaltering the principal direction of protein's motion, although it slows down protein dynamics so that the protein gets trapped in the homogeneous ensemble of conformations closer to the native state. Free energy landscape reveals higher population of native compared to intermediate and denatured states. Delphi results and calculation of the preferential interaction parameter demonstrate that this relative stabilization of the native state can be ascribed to be the consequence of favourable interactions of trehalose with side chains of certain loci on the protein surface encompassing polar flexible residues. Stability of protein results from the observed difference in binding affinity of trehalose for native and denatured states of protein. Our findings are at variance with the common conception of relative destabilization of the denatured state. Rather, we provide evidence for relative stabilization of the native state. This stabilization is due to interplay of protein-trehalose, water-trehalose, water-water, protein-water and trehalose-trehalose interactions.

Thermodynamic description of Hofmeister effects on the LCST of thermosensitive polymers.

Cosolvent effects on protein or polymer collapse transitions are typically discussed in terms of a two-state free energy change that is strictly linear in cosolute concentration. Here we investigate in detail the nonlinear thermodynamic changes of the collapse transition occurring at the lower critical solution temperature (LCST) of the role-model polymer poly(N-isopropylacrylamide) [PNIPAM] induced by Hofmeister salts. First, we establish an equation, based on the second-order expansion of the two-state free energy in concentration and temperature space, which excellently fits the experimental LCST curves and enables us to directly extract the corresponding thermodynamic parameters. Linear free energy changes, grounded on generic excluded-volume mechanisms, are indeed found for strongly hydrated kosmotropes. In contrast, for weakly hydrated chaotropes, we find significant nonlinear changes related to higher order thermodynamic derivatives of the preferential interaction parameter between salts and polymer. The observed non-monotonic behavior of the LCST can then be understood from a not yet recognized sign change of the preferential interaction parameter with salt concentration. Finally, we find that solute partitioning models can possibly predict the linear free energy changes for the kosmotropes, but fail for chaotropes. Our findings cast strong doubt on their general applicability to protein unfolding transitions induced by chaotropes.