Binding Interaction of Coumarin Derivative Daphnetin with Ovalbumin: Molecular Insights into the Complexation Process and Effects of Metal Ions and pH in the Binding and Antifibrillation Studies.

This study investigates the interaction between daphnetin and ovalbumin (OVA) as well as its potential to inhibit OVA fibrillation using both spectroscopic and computational analysis. A moderate binding affinity of 1 × 104 M-1 was observed between OVA and daphnetin, with a static quenched mechanism identified during the fluorescence quenching processes. Metal ions' (Cu2+ and Zn2+) presence led to an increase in the binding affinities of daphnetin toward OVA, mirroring a similar trend observed with the pH variation. Synchronous and 3D fluorescence studies indicated an increase in the polarity of the microenvironment surrounding the Trp residues during binding. Interestingly, circular dichroism and Fourier transform infrared studies showed a significant change in the secondary structure of OVA upon binding with daphnetin. The efficacy of daphnetin in inhibiting protein fibrillation was confirmed through thioflavin T and Congo Red binding assays along with fluorescence microscopic imaging analysis. The thermodynamic assessment showed positive ΔH° [+(29.34 ± 1.526) kJ mol-1] and ΔS° [+(181.726 ± 5.465) J mol-1] values, indicating the presence of the hydrophobic forces, while negative ΔG° signifies spontaneous binding interactions. These experimental findings were further correlated with computational analysis, revealing daphnetin dynamics within the binding site of OVA.

Interfacial Glucose to Regulate Hydrated Lipid Bilayer Properties: Influence of Concentrations.

A series of atomistic molecular dynamics (MD) simulations were carried out with a hydrated 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayer with the variation of glucose concentrations from 0 to 30 wt % in the presence of 0.3 M NaCl. The study suggested that although the thickness of the lipid bilayer dropped significantly with the increase in glucose concentration, it expanded laterally at high glucose levels due to the intercalation of glucose between the headgroups of adjacent lipids. We adopted the surface assessment via the grid evaluation method to compute the deviation of the bilayer's key structural features for the different amounts of glucose present. This suggested that the accumulation of glucose molecules near the headgroups influences the local lipid bilayer undulation and crimping of the lipid tails. We find that the area compressibility modulus increases with the glucose level, causing enhanced bilayer rigidity arising from the slow lateral diffusion of lipids. The restricted lipid motion at high glucose concentrations controls the sustainability of the curved bilayer surface. Calculations revealed that certain orientations of CO→ of interfacial glucose with the PN→ of lipid headgroups are preferred, which helps the glucose to form direct hydrogen bonds (HBs) with the lipid headgroups. Such lipid-glucose (LG) HBs relax slowly at low glucose concentrations and exhibit a higher lifetime, whereas fast structural relaxation of LG HBs with a shorter lifetime was noticed at a higher glucose level. In contrast, lipid-water (LW) HBs exhibited a higher lifetime at a higher glucose level, which gradually decreased with the glucose level lowering. The study interprets that the glucose concentration-driven LW and LG interactions are mutually inclusive. Our detailed analysis will exemplify small saccharide concentration-driven membrane stabilizing efficiency, which is, in general, helpful for drug delivery study.

Conformational preferences of heparan sulfate to recognize the CXCL8 dimer in aqueous medium: degree of sulfation and hydrogen bonds.

The sulfation pattern and epimerization of the long-chain sulfated polysaccharide heparan sulfate (HS) cause structural diversity and regulate various physiological and pathological processes when binding with proteins. In this work, we performed a series of molecular dynamics simulations of three variants of the octadecasaccharide HS with varying sulfation positions in aqueous medium in their free forms and in the presence of the chemokine CXCL8 dimer. The free energy of binding depicts the sulfation at the 6-O position of GlcNAc (HS6S), and both 3-O and 6-O positions of GlcNAc (HS3S6S) of HS variants are more likely to bind with the CXCL8 dimer than the triply sulfated HS2S3S6S, which is sulfated at the 2-O position of GlcUA additionally along with 3-O and 6-O positions of GlcNAc. Binding between HS and CXCL8 was driven by electrostatic and van der Waals interactions predominantly regardless of the sulfation pattern; however, unfavorable entropic contribution suppressed the interaction between HS and CXCL8. The contribution of different amino acid residues to the binding energetics suggested that basic amino acids line up the binding site of CXCL8. This study further acknowledges the role of interfacial water that is structured and bound with HS through hydrogen bonds, exhibiting differential hydrogen bond relaxation dynamics compared to when the HS molecules are free. Moreover, this study identifies that with the increase in sulfation, the HS-water hydrogen bond relaxation occurs faster with the complexation, while the reverse trend is followed in their free forms. Significant structural adaptation of the different sulfated HS molecules, as verified from the free energy landscapes generated from various reaction coordinates, root-mean-square-deviations, end-to-end distances, including ring pucker angles, dihedral flexibility, and the high conformational entropy cost arising from the glycosidic bonds, suggests that the different sulfated variants of HS undergo significant structural transformation to bind with CXCL8. The presence of a CXCL8 dimer imposes the bound forms of HS to adopt non-linear structures with skew-boat conformations. The atomistic details of the study would help in understanding the selectivity and conformational diversity, as well as the role of solvents in the recognition of CXCL8 by different sulfated variants of HS molecules.

Boron-containing fullerene-based salts with cyclic carbonate solvents as electrolytes for Li-ion batteries and beyond.

Replacement of carbon atoms by a heteroatom in fullerene is a promising route that enhances the electronic properties of fullerenes and results in hetero fullerene-based effective agents ensuring applications in vivid fields of the solar cell, cathode materials for batteries, etc. Towards the development of new electrolyte salts, attention has been paid to facilitating ion mobility in particular and moderate stability of the anions in addition. From the atomistic molecular dynamics simulation studies, for the first time, we uncover that the boron-containing hetero fullerene, C59B- anion-based LiC59B, and NaC59B salts in cyclic carbonate solvents can act as efficient electrolytes by improving the transport phenomenon of the metal ions in solution, importantly for Li+ and satisfactorily for Na+ as compared to their commonly used BF4- anion based salts. Additionally, our study revealed that apart from LiC59B, and NaC59B salts, C58B22- based MgC58B2 salt can facilitate the ionic conductivity of the electrolyte. The properties of the proposed electrolyte under an electric field and different temperatures were investigated. Some of the bulk properties of the used electrolytes to some extent were found to be improved in the presence of these salts. The first principle-based electrochemical calculations further justify the stability of the proposed anions. The initial investigation from the Reactive force-field (ReaxFF) based atomistic simulations study elucidates that LiC59B reduces the decomposition of the EC solvent compared to LiBF4 and facilitates solvent stability.

Understanding Conformational Properties and Role of Hydrogen Bonds in Glycosaminoglycans-Interleukin8 Complexes in Aqueous Medium by Molecular Dynamics Simulation.

Atomistic molecular dynamics simulations were performed under ambient conditions to explore the conformational features and binding affinities of hexameric glycosaminoglycans (GAGs) with chemokine Interleukin8 (IL8) in an aqueous medium. We tried to understand the role of hydrogen bonds (HBs) involving conserved water in mediating the interactions. The Luzar-Chandler model was adopted to study the kinetics of HB breaking and formation concerning different water-mediated HBs. The conformational flexibilities of bound GAGs are due to the flexible glycosidic linkages than the occasional/rare ring pucker conformation. The free energy landscape constructed with ϕ, and ψ, depicted that different conformational minima associated with the glycosidic linkage flexibility of the GAGs in bound states are separated by energy barriers. The binding affinities of IL8 towards GAGs are favored through the electrostatic and non-polar solvation interactions. 4-different types of conserved water were explored in the solvent-mediated binding of GAGs with IL8. The average lifetime of the IL8-GAG direct HB pairs was ∼ten times less than the IL8-GAG-shared water HBs. This is due to the rapid establishment of HB breaking and reformation kinetics involving water of a shared layer. We find that despite the highly negatively charged surface of GAGs, the IL8 surface populated by non-cationic amino acids could serve as a promising binding site in addition to the cationic surface of the protein.

Sulfation Effects of Chondroitin Sulfate to Bind a Chemokine in Aqueous Medium: Conformational Heterogeneity and Dynamics from Molecular Simulation.

The sulfation patterns and degree of sulfation of chondroitin sulfate (CS), an important class of glycosaminoglycans (GAG), and their interactions with chemokines are accountable for various diseases. To realize the underlying mechanism of such complex biological phenomena at a molecular level and their application in rational drug design, a study on conformations and dynamics of CSs is necessary. To explore this, in this study, we performed a series of atomistic molecular dynamics (MD) simulations with different sulfated variants of octadecasaccharide CS, like CS-C, CS-E, and CS-T, in their free forms and when bound to the protein chemokine CXCL8 dimer in an aqueous medium. The calculated binding free energy of CSs with the CXCL8 dimer is favorable, and the degree of sulfation favors the complexation process further with prominent hydrophobic and hydrogen-bonded interactions. We find that the recognition is associated with the configurational entropy loss of the CS molecules as calculated from the Gaussian mixture approach, which supports that the degree of sulfation regulates the process. Cluster analysis through the k-means algorithm and end-to-end distance measurement revealed that although the free CS molecules adopted linear conformations, the nonlinear conformations during binding with protein were noted. Adaptation of nonlinear forms in the bound forms is noteworthy for the less-sulfated CS-C and CS-E. Apart from favorable 4C1 conformations, the occasional appearance of skew-boat forms from the free-energy map of ring pucker for the GlcUA unit was observed, which remains unaffected by the sulfation. We find that during recognition, the average relaxation time of intra-CS and inter-CS-CXCL8 hydrogen bonds (HBs) is about a magnitude lesser than that of CS-water HBs, most prominent on the involvement of higher sulfated CS-T analogues. The translational motion of surrounded water molecules in CSs exhibited sublinear diffusion, and the degree of sublinearity increases around the heavily sulfated molecules due to the hindrance created by them as well as the presence of the chemokine and exhibited markedly slow heterogeneous diffusion.

Unraveling the origin of interactions of hydroxychloroquine with the receptor-binding domain of SARS-CoV-2 in aqueous medium.

Interactions of hydroxychloroquin (HCQ) with the receptor binding domain (RBD) of SARS-CoV-2 were studied from atomistic simulation and ONIOM techniques. The key-residues of RBD responsible for the human transmission are recognized to be blocked in a heterogeneous manner with the favorable formation of key-residue:HCQ (1:1) complex. Such heterogeneity in binding was identified to be governed by the differential life-time of the hydrogen bonded water network anchoring HCQ and the key-residues. The intermolecular proton transfer facilitates the most favorable Lys417:HCQ complexation. The study demonstrates that off-target bindings of HCQ need to be minimized to efficiently prevent the transmission of SARS-CoV-2.

Analyzing the driving forces of insulin stability in the basic amino acid solutions: A perspective from hydration dynamics.

Amino acids having basic side chains, as additives, are known to increase the stability of native-folded state of proteins, but their relative efficiency and the molecular mechanism are still controversial and obscure as well. In the present work, extensive atomistic molecular dynamics simulations were performed to investigate the hydration properties of aqueous solutions of concentrated arginine, histidine, and lysine and their comparative efficiency on regulating the conformational stability of the insulin monomer. We identified that in the aqueous solutions of the free amino acids, the nonuniform relaxation of amino acid-water hydrogen bonds was due to the entrapment of water molecules within the amino acid clusters formed in solutions. Insulin, when tested with these solutions, was found to show rigid conformations, relative to that in pure water. We observed that while the salt bridges formed by the lysine as an additive contributed more toward the direct interactions with insulin, the cation-π was more prominent for the insulin-arginine interactions. Importantly, it was observed that the preferentially more excluded arginine, compared to histidine and lysine from the insulin surface, enriches the hydration layer of the protein. Our study reveals that the loss of configurational entropy of insulin in arginine solution, as compared to that in pure water, is more as compared to the entropy loss in the other two amino acid solutions, which, moreover, was found to be due to the presence of motionally bound less entropic hydration water of insulin in arginine solution than in histidine or lysine solution.

Insights into the Sensitivity of Arginine Concentration to Preserve the Folded Form of Insulin Monomer under Thermal Stress.

Arginine, although popularly known as aggregation suppressor additive, has been found to quench proteins' structure and function by destabilizing their conformations. Driven by such controversial evidence, in this work we performed a series of atomistic molecular dynamics simulations of insulin monomer, a biologically active hormone protein, in arginine solution of varying concentrations (0.5, 1, and 2 M) at ambient and elevated temperature (400 K) to explore the arginine concentration driven structure-based stability of the protein. Our study reveals that the flexibility of the protein's structure is dependent on the arginine concentration, and among all the used solutions, 2 M arginine, a "neutral crowder" that mimics the cellular environment, can preserve the native folded form of the protein at ambient temperature in an excellent manner. Further, while the protein unfolds at 400 K in pure water, this solution worked satisfactorily to preserve the protein's folded conformation more firmly than the other solutions. The replica-exchange MD of insulin in 2 M arginine solution further supports the fact. In this aspect an important issue in molecular pharmacology is to identify and recognize the physical origin of the stability of a protein, i.e, in this case, how arginine directs the conformational flexibility of the protein and preserves its native folded form. We identified that the exclusion of arginine from the protein surface increases the local structuration of water around the protein, thereby preserving its "biological water" layer, and makes the protein more hydrated at 2 M concentration as compared to the other arginine solutions. Additionally, our microscopic investigation on the interactions of the protein-solvation layer revealed that the structural heterogeneity of the protein surface, arising from the differential physicochemical nature of the amino acid residues, controls the favorable formation of sluggish water-arginine mixed solvation layer at higher arginine concentration that helps the protein to maintain its structural rigidity. Importantly, apart from the protein-solvent hydrogen-bonding interactions, the anion-pi interactions, established between the carboxyl group of arginine and the aromatic amino acid residues of insulin, were recognized to facilitate the protein to maintain its native folded form at the experimental temperatures.

Superhalogens as Building Blocks of Super Lewis Acids.

Lewis acids play an important role in synthetic chemistry. Using first-principle calculations on some newly designed molecules containing boron and organic heterocyclic superhalogen ligands, we show that the acid strength depends on the charge of the central atom as well as on the ligands attached to it. In particular, the strength of the Lewis acid increases with increasing electron withdrawing power of the ligand. With this insight, we highlight the importance of superhalogen-based ligands in the design of strong Lewis acids. Calculated fluoride ion affinity (FIA) values of B[C2 BNO(CN)3 ]3 and B[C2 BNS(CN)3 ]3 show that these are super Lewis acids.

A New Class of Superhalogen Based Anion Receptor in Li-Ion Battery Electrolytes.

In the search for new additives (anion receptors) in Li-ion battery electrolytes especially for LiPF6 and LiClO4, we have theoretically designed boron-based complexes by coupling with different heterocyclic ligands. The validation of the formation of modeled compounds involves reproduction of available experimentally reported absolute magnetic shielding and chemical shift values for different boron complexes. As compared to the commonly used tris(pentafluorophenyl) borane, our designed compounds suggest that the complexes like B[C2HBNO(CN)2]3, B[C2HBNS(CN)2]3, and B[C4H3BN(CN)2]3 are promising additives.

Lysozyme-luteolin binding: molecular insights into the complexation process and the inhibitory effects of luteolin towards protein modification.

In the proposed work, the complexation of bioactive flavonoid luteolin with hen egg white lysozyme (HEWL) along with its inhibitory influence on HEWL modification has been explored with the help of multi-spectroscopic and computational methods. The binding affinity has been observed to be moderate in nature (in the order of 104 M-1) and the static quenching mechanism was found to be involved in the fluorescence quenching process. The binding constant (Kb) shows a progressive increase with the increase in temperature from (4.075 ± 0.046 × 104 M-1) at 293 K to (6.962 ± 0.024 × 104 M-1) at 313 K under experimental conditions. Spectroscopic measurements along with molecular docking calculations suggest that Trp62 is involved in the binding site of luteolin within the geometry of HEWL. The positive changes in enthalpy (ΔH = +19.99 ± 0.65 kJ mol-1) as well as entropy (ΔS = +156.28 ± 2.00 J K-1 mol-1) are indicative of the presence of hydrophobic forces that stabilize the HEWL-luteolin complex. The micro-environment around the Trp residues showed an increase in hydrophobicity as indicated by synchronous fluorescence (SFS), three dimensional fluorescence (3D) and red edge excitation (REES) studies. The % α-helix of HEWL showed a marked reduction upon binding with luteolin as indicated by circular dichroism (CD) and Fourier-transform infrared spectroscopy (FTIR) studies. Moreover, luteolin is situated at a distance of 4.275 ± 0.004 nm from the binding site as indicated by FRET theory, and the rate of energy transfer kET (0.063 ± 0.004 ns-1) has been observed to be faster than the donor decay rate (1/τD = 0.606 ns-1), which is indicative of the non-radiative energy transfer during complexation. Leaving aside the binding study, luteolin showed promising inhibitory effects towards the d-ribose mediated glycation of HEWL as well as towards HEWL fibrillation as studied by fluorescence emission and imaging studies. Excellent correlation with the experimental observations as well as precise location and dynamics of luteolin within the binding site has been obtained from molecular docking and molecular dynamics simulation studies.

Can 2,2,2-trifluoroethanol be an efficient protein denaturant than methanol and ethanol under thermal stress?

Monohydric alcohols, such as methanol (MEH), ethanol (ETH) and 2,2,2-trifluoroethanol (TFE), have significant effects on biological processes including the protein folding-unfolding phenomenon. Among the several monohydric alcohols, TFE, a fluorine-substituted alcohol, is known to induce a helical structure in proteins. In this work, we report the heterogeneous unfolding phenomenon of a small protein Chymotrypsin Inhibitor 2 in various concentrations of methanol, ethanol and TFE solutions by performing atomistic molecular dynamics simulation studies. Our study reveals that the unfolding phenomenon of CI2 under thermal stress majorly depends on the concentration and the nature of the alcohol. The presence of alcohols in general has been noted to accelerate the unfolding process compared to pure water and TFE, among them all, has been found to speed up the unfolding time scale at low concentrations. The molecular contact frequency between protein and alcohol follows the trend, MEH < ETH < TFE at low concentrations, whereas the trend becomes MEH ∼ ETH > TFE at more concentrated solutions. The differential water-mediated and self-clustering phenomena of alcohols, diverse protein-alcohol hydrogen bond strengths and the concentration dependent restricted inhomogeneous protein-water as well as protein-alcohol hydrogen bond dynamics suggest that TFE, a well known α-helix stabilizer, could be a good competitor among its class of denaturants.

Conformational disorder and solvation properties of the key-residues of a protein in water-ethanol mixed solutions.

A small number of key-residues in a protein sequence play vital roles in the function, stability, and folding of the protein. The nonuniform conformational disorder of a small protein Chymotrypsin Inhibitor 2 (CI2) and its secondary segments has been quantified in the ethanol governed temperature induced unfolding process by estimating its change in configurational entropy in several water-ethanol mixed solutions. Such calculations further assist us in identifying the key-residues, from where the unfolding of the protein was initiated. Our findings match well with the reported experimental results. We then make an attempt to explore the properties of the solvent water and ethanol around the key-residues of the protein in its folded and unfolded forms at ambient temperature to identify the individual role of ethanol and water in the protein unfolding. We find that the key-residues of the unfolded protein are in good contact with both water and ethanol as compared to those of the folded protein. In the presence of ethanol, water molecules are noticed to form a rigid structurally bound solvation layer around the key-residues of the protein, irrespective of its conformational state. The restricted translational motion and prominent caging effect of the water and ethanol molecules present around the key-residues of the unfolded protein are a signature of the existence of a rigid mixed water-ethanol layer as compared to that around the folded protein. Furthermore, comparable restricted structural relaxation of the key-residue-water and key-residue-ethanol hydrogen bonds in the unfolded protein as compared to that in the folded one implies that the formation of a strong long-lived hydrogen bonding environment nourishes the unfolding process. We believe that our findings will shed light to several co-solvent governed unfolding processes of a protein in general.

Residue Specific Interaction of an Unfolded Protein with Solvents in Mixed Water-Ethanol Solutions: A Combined Molecular Dynamics and ONIOM Study.

The molecular mechanism of ethanol governed unfolding of an enzymatic protein, chymotrypsin inhibitor 2 (CI2), in water-ethanol mixed solutions has been studied by using combined molecular dynamics simulations and ONIOM study. The residue specific solvation of the unfolded protein and the interactions between the individual amino acid residues of the protein with ethanol as well as water have been investigated. The results are compared with that obtained from the folded state of the protein. Further, emphasis has been given to explore the residue's preferential site of attraction toward the nature of the solvents. The heterogeneous structuring of water and ethanol around the hydrophobic and hydrophilic surfaces of the protein is found to correlate well with their available surface areas to the solvents. Both hydrophobic and hydrophilic interactions are found to have important contributions in rupturing protein's secondary structural segments. Further, residue-water as well as residue-ethanol binding energies show significant involvement of the hydrogen bonding environment in the unfolding process; particularly, residue-water hydrogen bonds are found to play an indispensable role.

Insights into the Mechanism of Ground and Excited State Double Proton Transfer Reaction in Formic Acid Dimer.

The mechanism of ground and excited state double proton transfer reaction in formic acid dimer has been analyzed with the help of reaction force and the reaction electronic flux. The separation of reaction electronic flux in terms of electronic activity and reactivity, NBO, and dual descriptor lends additional support for the mechanism. Interestingly we found that the ground state double proton transfer mechanism is concerted synchronic, whereas the excited state double proton transfer is concerted asynchronic in nature.

Delineating the conformational flexibility of trisaccharides from NMR spectroscopy experiments and computer simulations.

The conformation of saccharides in solution is challenging to characterize in the context of a single well-defined three-dimensional structure. Instead, they are better represented by an ensemble of conformations associated with their structural diversity and flexibility. In this study, we delineate the conformational heterogeneity of five trisaccharides via a combination of experimental and computational techniques. Experimental NMR measurements target conformationally sensitive parameters, including J couplings and effective distances around the glycosidic linkages, while the computational simulations apply the well-calibrated additive CHARMM carbohydrate force field in combination with efficient enhanced sampling molecular dynamics simulation methods. Analysis of conformational heterogeneity is performed based on sampling of discreet states as defined by dihedral angles, on root-mean-square differences of Cartesian coordinates and on the extent of volume sampled. Conformational clustering, based on the glycosidic linkage dihedral angles, shows that accounting for the full range of sampled conformations is required to reproduce the experimental data, emphasizing the utility of the molecular simulations in obtaining an atomic detailed description of the conformational properties of the saccharides. Results show the presence of differential conformational preferences as a function of primary sequence and glycosidic linkage types. Significant differences in conformational ensembles associated with the anomeric configuration of a single glycosidic linkage reinforce the impact of such changes on the conformational properties of carbohydrates. The present structural insights of the studied trisaccharides represent a foundation for understanding the range of conformations adopted in larger oligosaccharides and how these molecules encode their conformational heterogeneity into the monosaccharide sequence.

Effect of ethanol concentrations on temperature driven structural changes of chymotrypsin inhibitor 2.

A series of atomistic molecular dynamics (MD) simulations of a small enzymatic protein Chymotrypsin Inhibitor 2 (CI2) in water-ethanol mixed solutions were carried out to explore the underlying mechanism of ethanol driven conformational changes of the protein. Efforts have been made to probe the influence of ethanol concentrations ranging from 0% to 75% (v/v) at ambient condition (300 K (T1)) and at elevated temperatures (375 K (T2) and 450 K (T3)) to investigate the temperature induced conformational changes of the protein further. Our study showed that the effect of varying ethanol concentrations on protein's structure is almost insignificant at T1 and T2 temperatures whereas at T3 temperature, partial unfolding of CI2 in 10% ethanol solution followed by full unfolding of the protein at ethanol concentrations above 25% occurs. However, interestingly, at T3 temperature CI2's native structure was found to be retained in pure water (0% ethanol solution) indicating that the cosolvent ethanol do play an important role in thermal denaturation of CI2. Such observations were quantified in the light of root-mean-square deviations (RMSDs) and radius of gyration. Although higher RMSD values of ß-sheet over α-helix indicate complete destruction of the ß-structure of CI2 at high ethanol concentrations, the associated time scale showed that the faster melting of α-helix happens over ß-sheet. Around 60%-80% of initial native contacts of the protein were found broken with the separation of hydrophobic core consisting eleven residues at ethanol concentrations greater than 25%. This leads protein to expand with the increase in solvent accessible surface area. The interactions between protein and solvent molecules showed that protein's solvation shell preferred to accommodate ethanol molecules as compared to water thereby excluded water molecules from CI2's surface. Further, concentration dependent differential self-aggregation behavior of ethanol is likely to regulate the replacement of relatively fast diffused water by low diffused ethanol molecules from protein's surface during the unfolding process.

In silico studies of the properties of water hydrating a small protein.

Atomistic molecular dynamics simulation of an aqueous solution of the small protein HP-36 has been carried out with explicit solvent at room temperature. Efforts have been made to explore the influence of the protein on the relative packing and ordering of water molecules around its secondary structures, namely, three α-helices. The calculations reveal that the inhomogeneous water ordering and density distributions around the helices are correlated with their relative hydrophobicity. Importantly, we have identified the existence of a narrow relatively dehydrated region containing randomly organized "quasi-free" water molecules beyond the first layer of "bound" waters at the protein surface. These water molecules with relatively weaker binding energies form the transition state separating the "bound" and "free" water molecules at the interface. Further, increased contribution of solid-like caging motions of water molecules around the protein is found to be responsible for reduced fluidity of the hydration layer. Interestingly, we notice that the hydration layer of helix-3 is more fluidic with relatively higher entropy as compared to the hydration layers of the other two helical segments. Such characteristics of helix-3 hydration layer correlate well with the activity of HP-36, as helix-3 contains the active site of the protein.

Conformational Features and Hydration Dynamics of Proteins in Cosolvents: A Perspective from Computational Approaches.

The importance of solvent in stabilizing protein structures has long been recognized. Water is the common solvent for proteins, and hydration is elemental in governing protein stability, flexibility, and function through various interactions. The addition of small organic molecules known as cosolvents may deploy stabilization (folding) or destabilization (unfolding) effects on native protein conformations. Despite exhaustive literature, the molecular mechanism by which cosolvents regulate protein conformations and dynamics is controversial. Specifically, the cosolvent behavior has been unpredictable with the nature and concentrations that lead to protein stabilizing/destabilizing effects as it changes in water content near the vicinity of proteins. With the massive development of computational resources, advancement of computational methods, and the availability of numerous experimental techniques, various theoretical and computational studies of proteins in a mixture of solvents have been instigated. The growing interest in such studies has been to unravel the underlying mechanism of protein folding and cosolvent/solvent-protein interactions that have significant implications in biomedical and biotechnological applications. In this mini-review, apart from the brief overview of important theories and force-field model-based cosolvent effects on proteins, we present the current state of knowledge and recent advances in the field to describe cosolvent-guided conformational features of proteins and hydration dynamics from computational approaches. The mini-review further explains the mechanistic details of protein stability in various popularly used cosolvents, including limitations of present studies and future outlooks. The counteracting effects of cosolvent on the proteins in the mixture of stabilizing and destabilizing cosolvents are also presented and discussed.