Loss Of Conformational Entropy Research Articles

Engineering de novo disulfide bond in bacterial α-type carbonic anhydrase for thermostable carbon sequestration.

Exploiting carbonic anhydrase (CA), an enzyme that rapidly catalyzes carbon dioxide hydration, is an attractive biomimetic route for carbon sequestration due to its environmental compatibility and potential economic viability. However, the industrial applications of CA are strongly hampered by the unstable nature of enzymes. In this work, we introduced in silico designed, de novo disulfide bond in a bacterial α-type CA to enhance thermostability. Three variants were selected and expressed in Escherichia coli with an additional disulfide bridge. One of the variants showed great enhancement in terms of both kinetic and thermodynamic stabilities. This improvement could be attributed to the loss of conformational entropy of the unfolded state, showing increased rigidity. The variant showed an upward-shifted optimal temperature and appeared to be thermoactivated, which compensated for the lowered activity at 25 °C. Collectively, the variant constructed by the rapid and effective de novo disulfide engineering can be used as an efficient biocatalyst for carbon sequestration under high temperature conditions.

Acetylation of Surface Lysine Groups of a Protein Alters the Organization and Composition of Its Crystal Contacts.

This paper uses crystals of bovine carbonic anhydrase (CA) and its acetylated variant to examine (i) how a large negative formal charge can be accommodated in protein-protein interfaces, (ii) why lysine residues are often excluded from them, and (iii) how changes in the surface charge of a protein can alter the structure and organization of protein-protein interfaces. It demonstrates that acetylation of lysine residues on the surface of CA increases the participation of polar residues (particularly acetylated lysine) in protein-protein interfaces, and decreases the participation of nonpolar residues in those interfaces. Negatively charged residues are accommodated in protein-protein interfaces via (i) hydrogen bonds or van der Waals interactions with polar residues or (ii) salt bridges with other charged residues. The participation of acetylated lysine in protein-protein interfaces suggests that unacetylated lysine tends to be excluded from interfaces because of its positive charge, and not because of a loss in conformational entropy. Results also indicate that crystal contacts in acetylated CA become less constrained geometrically and, as a result, more closely packed (i.e., more tightly clustered spatially) than those of native CA. This study demonstrates a physical-organic approach-and a well-defined model system-for studying the role of charges in protein-protein interactions.

A helical peptide confined in metal-organic frameworks: Microscopic insight from molecular simulation

We report a molecular simulation study to investigate a helical peptide (β-adrenoceptor) confined in two metal-organic frameworks (MOFs) namely IRMOF-74-II and -III. Reversible structural transformation between the major (α-helix and 310-helix) and minor structures (bend, turn and coil) of the confined peptide is observed. In IRMOF-74-II, the peptide is over-confined and the helicity is lower compared to that in bulk water; however, the helicity is enhanced upon moderately confined in IRMOF-74-III. As attributed to the loss of conformational entropy, the thermal fluctuations of the peptide in both MOFs are smaller than in bulk water. The hydrophobic solvent-accessible surface area of the peptide, in contrast to the hydrophilic counterpart, is largely reduced upon confined in both MOFs because of the favorable interactions between hydrophobic residues and MOFs. This study provides microscopic insight into β-adrenoceptor peptide confined in MOFs and might be useful toward the rational design of nano-carriers for peptides and other biomolecules.

Statistical Thermodynamics for Actin-Myosin Binding: The Crucial Importance of Hydration Effects

Actomyosin is an important molecular motor, and the binding of actin and myosin is an essential research target in biophysics. Nevertheless, the physical factors driving or opposing the binding are still unclear. Here, we investigate the role of water in actin-myosin binding using the most reliable statistical-mechanical method currently available for assessing biomolecules immersed in water. This method is characterized as follows: water is treated not as a dielectric continuum but as an ensemble of molecules; the polyatomic structures of proteins are taken into consideration; and the binding free energy is decomposed into physically insightful entropic and energetic components by accounting for the hydration effect to its full extent. We find that the actin-myosin binding brings large gains of electrostatic and Lennard-Jones attractive interactions. However, these gains are accompanied by even larger losses of actin-water and myosin-water electrostatic and LJ attractive interactions. Although roughly half of the energy increase due to the losses is cancelled out by the energy decrease arising from structural reorganization of the water released upon binding, the remaining energy increase is still larger than the energy decrease brought by the gains mentioned above. Hence, the net change in system energy is positive, which opposes binding. Importantly, the binding is driven by a large gain of configurational entropy of water, which surpasses the positive change in system energy and the conformational entropy loss occurring for actin and myosin. The principal physical origin of the large water-entropy gain is as follows: the actin-myosin interface is closely packed with the achievement of high shape complementarity on the atomic level, leading to a large increase in the total volume available to the translational displacement of water molecules in the system and a resultant reduction of water crowding (i.e., entropic correlations among water molecules).

Flattening Process of Polymer Chains Irreversibly Adsorbed on a Solid.

We report the structural relaxation process of irreversibly adsorbed polymer chains via thermal annealing that lie flat on a solid ("flattened chains"). Amorphous polystyrene and quartz, which together constitute a weakly attractive system, was used as a model where the local chain conformations of the flattened chains were investigated by sum frequency generation spectroscopy (SFG). Two different film preparation processes (i.e., spin coating and dip coating methods) were utilized to create different initial chain conformations. The spin-coated and dip-coated PS thin films were annealed at a temperature far above the bulk glass transition temperature to reach the "quasiequilibrium" state and subsequently rinsed with chloroform to uncover the buried flattened chains. The SFG results revealed that the backbone chains (constituted of CH and CH2 groups) of the flattened PS chains preferentially orient to the weakly interactive substrate surface via thermal annealing regardless of the initial chain conformations, while the orientation of the phenyl rings becomes randomized. We postulate that increasing the number of surface-segmental contacts (i.e., enthalpic gain) is the driving force for the flattening process of the polymer chains, even onto a weakly interactive solid to overcome the conformational entropy loss in the total free energy.

Lattice model of oligonucleotide hybridization in solution. II. Specificity and cooperativity.

Because oligonucleotides are short sequences of nucleic acid bases, their association in solution with complementary strands (hybridization) is often seen to conform to a simple two-state model. However, experimental evidence suggests that, despite their short length, oligonucleotides may hybridize through multiple states involving intermediates. We investigate whether these apparently contradictory scenarios are possible by imposing different levels of sequence specificity on a lattice model of oligonucleotides in solution, which we introduced in Part I [J. C. Araque et al., J. Chem. Phys. 134, 165103 (2011)]. We find that both multiple-intermediate (weakly cooperative) and two-state (strongly cooperative) transitions are possible and that these are directly linked to the level of sequence specificity. Sequences with low specificity hybridize (base-by-base) by way of multiple stable intermediates with increasing number of paired bases. Such intermediate states are weakly cooperative because the energetic gain from adding an additional base pair is outweighed by the conformational entropy loss. Instead, sequences with high specificity hybridize through multiple metastable intermediates which easily bridge the configurational and energetic gaps between single- and double-stranded states. These metastable intermediates interconvert with minimal loss of conformational entropy leading to a strongly cooperative hybridization. The possibility of both scenarios, multiple- and two-states, is therefore encoded in the specificity of the sequence which in turn defines the level of cooperativity.

Essential Roles of Water in Actin-Myosin Binding

Though the binding of actin and myosin is one of the most important biological self-assembly processes, its mechanism is still controversial. In a prevailing view, which claims that the binding is driven by the actin-myosin electrostatic attractive interaction, the effect of surrounding water molecules (i.e., hydration effect) is not suitably taken into account. Here we investigate the water roles in the binding using a statistical-mechanical approach combined with molecular models for water. The hydration energies and entropies of actin, myosin, and their complex are calculated by the three-dimensional reference interaction site model theory and by the angle-dependent integral equation theory combined with our morphometric approach, respectively. In this theory, the changes in thermodynamic quantities upon the binding can be decomposed into physically insightful components. The actin-myosin binding brings a large gain of their electrostatic attractive interaction. However, it accompanies an even larger loss of actin-water and myosin-water electrostatic attractive interactions. The net change in the electrostatic component of system energy is positive. We find that the binding is driven by a large gain of configurational entropy of water which should surpass the positive change in system energy and the conformational-entropy loss occurring for actin and myosin. The physical origin of the large water-entropy gain is as follows: The actin-myosin interface is closely packed with the achievement of high shape complementarity on the atomic level, leading to a large increase in the total volume available to the translational displacement of water molecules in the system.

Effect of backbone chemistry on hybridization thermodynamics of oligonucleic acids: a coarse-grained molecular dynamics simulation study.

In this paper we study how varying oligonucleic acid backbone chemistry affects the hybridization/melting thermodynamics of oligonucleic acids. We first describe the coarse-grained (CG) model with tunable parameters that we developed to enable the study of both naturally occurring oligonucleic acids, such as DNA, and their chemically-modified analogues, such as peptide nucleic acids (PNAs) and locked nucleic acids (LNAs). The DNA melting curves obtained using such a CG model and molecular dynamics simulations in an implicit solvent and with explicit ions match with the melting curves obtained using the empirical nearest-neighbor models. We use these CG simulations to then elucidate the effect of backbone flexibility, charge, and nucleobase spacing along the backbone on the melting curves, potential energy and conformational entropy change upon hybridization and base-pair hydrogen bond residence time. We find that increasing backbone flexibility decreases duplex thermal stability and melting temperature mainly due to increased conformational entropy loss upon hybridization. Removing charges from the backbone enhances duplex thermal stability due to the elimination of electrostatic repulsion and as a result a larger energetic gain upon hybridization. Lastly, increasing nucleobase spacing decreases duplex thermal stability due to decreasing stacking interactions that are important for duplex stability.

On the physics of thermal-stability changes upon mutations of a protein.

It is of great interest from both scientific and practical viewpoints to theoretically predict the thermal-stability changes upon mutations of a protein. However, such a prediction is an intricate task. Up to now, significantly many approaches for the prediction have been reported in the literature. They always include parameters which are adjusted so that the prediction results can be best fitted to the experimental data for a sufficiently large set of proteins and mutations. The inclusion is necessitated to achieve satisfactorily high prediction performance. A problem is that the resulting values of the parameters are often physically meaningless, and the physicochemical factors governing the thermal-stability changes upon mutations are rather ambiguous. Here, we develop a new measure of the thermal stability. Protein folding is accompanied by a large gain of water entropy (the entropic excluded-volume (EV) effect), loss of protein conformational entropy, and increase in enthalpy. The enthalpy increase originates primarily from the following: The energy increase due to the break of protein-water hydrogen bonds (HBs) upon folding cannot completely be cancelled out by the energy decrease brought by the formation of protein intramolecular HBs. We develop the measure on the basis of only these three factors and apply it to the prediction of the thermal-stability changes upon mutations. As a consequence, an approach toward the prediction is obtained. It is distinguished from the previously reported approaches in the following respects: The parameters adjusted in the manner mentioned above are not employed at all, and the entropic EV effect, which is ascribed to the translational displacement of water molecules coexisting with the protein in the system, is fully taken into account using a molecular model for water. Our approach is compared with one of the most popular approaches, FOLD-X, in terms of the prediction performance not only for single mutations but also for double, triple, and higher-fold (up to sevenfold) mutations. It is shown that on the whole our approach and FOLD-X exhibit almost the same performance despite that the latter uses the adjusting parameters. For multiple mutations, however, our approach is far superior to FOLD-X. Five multiple mutations for staphylococcal nuclease lead to highly enhanced stabilities, but we find that this high enhancement arises from the entropic EV effect. The neglect of this effect in FOLD-X is a principal reason for its ill success. A conclusion is that the three factors mentioned above play essential roles in elucidating the thermal-stability changes upon mutations.

An integrative structure-based framework for predicting biological effects mediated by antipeptide antibodies

A general framework is presented for predicting quantitative biological effects mediated by antipeptide antibodies, primarily on the basis of antigen structure (possibly featuring intrinsic disorder) analyzed to estimate epitope–paratope binding affinities, which in turn is considered within the context of dose–response relationships as regards antibody concentration. This is illustrated mainly using an approach based on protein structural energetics, whereby expected amounts of solvent-accessible surface area buried upon epitope–paratope binding are related to the corresponding binding affinity, which is estimated from putative B-cell epitope structure with implicit treatment of paratope structure, for antipeptide antibodies either reacting with peptides or cross-reacting with cognate protein antigens. Key methods described are implemented in SAPPHIRE/SUITE (Structural-energetic Analysis Program for Predicting Humoral Immune Response Epitopes/SAPPHIRE User Interface Tool Ensemble; publicly accessible via http://freeshell.de/~badong/suite.htm). Representative results thus obtained are compared with published experimental data on binding affinities and quantitative biological effects, with special attention to loss of paratope sidechain conformational entropy (neglected in previous analyses) and in light of key in-vivo constraints on antigen-antibody binding affinity and antibody-mediated effects. Implications for further refinement of B-cell epitope prediction methods are discussed as regards envisioned biomedical applications including the development of prophylactic and therapeutic antibodies, peptide-based vaccines and immunodiagnostics.

On the effect of hydrostatic pressure on the conformational stability of globular proteins.

The model developed for cold denaturation (Graziano, PCCP 2010, 12, 14245-14252) is extended to rationalize the dependence of protein conformational stability upon hydrostatic pressure, at room temperature. A pressure- volume work is associated with the process of cavity creation for the need to enlarge the liquid volume against hydrostatic pressure. This contribution destabilizes the native state that has a molecular volume slightly larger than the denatured state due to voids existing in the protein core. Therefore, there is a hydrostatic pressure value at which the pressure-volume contribution plus the conformational entropy loss of the polypeptide chain are able to overwhelm the stabilizing gain in translational entropy of water molecules, due to the decrease in water accessible surface area upon folding, causing denaturation.

Permeability of small molecules through vapor deposited polymer membranes

ABSTRACTFree‐standing, thin films of poly(2‐hydroxyethyl methacrylate), p(HEMA), membranes deposited by initiated chemical vapor deposition (iCVD) technique are presented. Systematic studies of permeation of model dye molecules through these free‐standing thin films are performed and effects of process parameters on the permeability and selectivity of the membranes synthesized using an all‐dry process are demonstrated for the first time. Permeation studies of model dye molecules show that the diffusion of large dye molecules through the membrane is faster than the smaller dye molecules, due to loss of conformation entropy. Mesh sizes of the membranes are controlled by tuning the crosslink ratio of the polymer and diffusion coefficients of alizarin yellow model dye molecules are obtained as 0.44 × 10−7 and 1.71 × 10−7 cm2/s for high and low crosslinked membranes, respectively. Selectivity of the membranes is also demonstrated by the permeation studies of dyes with similar sizes but different polarities and the permeability coefficient of the hydrophilic dye is observed to be 30 times larger than that of the hydrophobic dye of similar size. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015, 132, 42453.

Effects of Chain Rigidity on the Adsorption of a Polyelectrolyte Chain on Mixed Lipid Monolayer: A Monte Carlo Study

We apply Monte Carlo simulation to explore the adsorption of a positively charged polyelectrolyte on a lipid monolayer membrane, composed of electronically neutral, monovalent anionic and mulvitalent anionic phospholipids. We systematically assess the influence of various factors, including the intrinsic rigidity of the polyelectrolyte chain, the bead charge density of the polyelectrolyte, and the ionic strength of the saline solution, on the interfacial structural properties of the polyelectrolyte/monolayer complex. The enhancement of the polyelectrolyte chain intrinsic rigidity reduces the polyelectrolyte conformational entropy loss and the energy gains in electrostatic interaction, but elevates the segregated anionic lipid demixing entropy loss. This energy-entropy competition results in a nonmonotonic dependence of the polyelectrolyte/monolayer association strength on the degree of chain rigidity. The semiflexible polyelectrolyte, i.e., the one with an intermediate degree of chain rigidity, is shown to associate onto the ternary membane below a higher critical ionic concentration. In this ionic concentration regime, the semiflexible polyelectrolyte binds onto the monolayer more firmly than the pancake-like flexible one and exhibits a stretched conformation. When the chain is very rigid, the polyelectrolyte with bead charge density Zb = +1 exhibits a larger tail and tends to dissociate from the membrane, whereas the one with Zb= +2 can still bind onto the membrane in a bridge-like conformation. Our results imply that chain intrinsic rigidity serves as an efficient molecular factor for tailoring the adsorption/desorption transition and interfacial structure of the polyelectrolyte/monolayer complex.

Structural basis and energy landscape of apigenin-induced cancer cell apoptosis mechanism in PI3K/Akt pathway

This study attempted to elucidate how apigenin (AGI) interferes with the PI3K/Akt pathway to fulfill its anti-tumour activity by anchoring on the ATP-binding pocket of PI3K and PDK1. The structural basis and energetic property of AGI and ATP binding to human PDK1 and PI3K isoforms α, β, γ and δ were investigated in detail with homology modelling and molecular docking. Free binding energy calculations and molecular dynamics simulations revealed that AGI can cause less conformational entropy loss than ATP upon the binding and possess the same level of binding stability with that of ATP. Combining ADMET prediction, AGI was evaluated as a strong ATP-competitive inhibitor with good pharmacokinetics profile.

The cis conformation of proline leads to weaker binding of a p53 peptide to MDM2 compared to trans

The cis and trans conformations of the Xaa–Pro (Xaa: any amino acid) peptide bond are thermodynamically stable while other peptide bonds strongly prefer trans. The effect of proline cis–trans isomerization on protein binding has not been thoroughly investigated. In this study, computer simulations were used to calculate the absolute binding affinity for a p53 peptide (residues 17–29) to MDM2 for both cis and trans isomers of the p53 proline in position 27. Results show that the cis isomer of p53(17–29) binds more weakly to MDM2 than the trans isomer, and that this is primarily due to the difference in the free energy cost associated with the loss of conformational entropy of p53(17–29) when it binds to MDM2. The population of cis p53(17–29) was estimated to be 0.8% of the total population in the bound state. The stronger binding of trans p53(17–29) to MDM2 compared to cis may leave a minimal level of p53 available to respond to cellular stress. This study demonstrates that it is feasible to estimate the absolute binding affinity for an intrinsically disordered protein fragment binding to an ordered protein that are in good agreement with experimental results.

Disorder-to-order transition of MAGI-1 PDZ1 C-terminal extension upon peptide binding: thermodynamic and dynamic insights.

PDZ domains are highly abundant protein-protein interaction modules commonly found in multidomain scaffold proteins. The PDZ1 domain of MAGI-1, a protein present at cellular tight junctions that contains six PDZ domains, is targeted by the E6 oncoprotein of the high-risk human papilloma virus. Thermodynamic and dynamic studies using complementary isothermal titration calorimetry and nuclear magnetic resonance (NMR) (15)N heteronuclear relaxation measurements were conducted at different temperatures to decipher the molecular mechanism of this interaction. Binding of E6 peptides to the MAGI-1 PDZ1 domain is accompanied by an unusually large and negative change in heat capacity (ΔC(p)) that is attributed to a disorder-to-order transition of the C-terminal extension of the PDZ1 domain upon E6 binding. Analysis of temperature-dependent thermodynamic parameters and (15)N NMR relaxation data of a PDZ1 mutant in which this disorder-to-order transition was abolished allows the unusual thermodynamic signature of E6 binding to be correlated to local folding of the PDZ1 C-terminal extension. Comparison of the exchange contributions observed for wild-type and mutant proteins explains how variation in the solvent-exposed area may compensate for the loss of conformational entropy and further designates a distinct set of a few residues that mediate this local folding phenomena.

Glass Transition Temperature of Cyclic Stars.

Cyclic homo- and diblock copolymers with different topologies were synthesized using a combination of "living" radical polymerization and "click" coupling reactions. The topologies included 2- and 3-arm stars, with the arms consisting of either cyclic or linear polystyrene. In addition, a diblock consisting of a cyclic polystyrene and a cyclic poly(acrylic acid) was also made. The topologies by imposing topological constraints due to the presence of cyclic polymers and branch points had a marked influence on the glass transition temperature (Tg). It was found that for the polystyrene topology series, the Tg increased above the glass transition temperature at infinite molecular weight for a linear chain (i.e., Tg∞) and correlated to the more compact nature of cyclic polymers. For the cyclic diblock of polystyrene and poly(acrylic acid), the Tg increased significantly due to separation of the blocks into their pure phases. This resulted in significant stretching of the chains and thus loss of conformation entropy.

Loss of conformational entropy in protein folding calculated using realistic ensembles and its implications for NMR-based calculations.

The loss of conformational entropy is a major contribution in the thermodynamics of protein folding. However, accurate determination of the quantity has proven challenging. We calculate this loss using molecular dynamic simulations of both the native protein and a realistic denatured state ensemble. For ubiquitin, the total change in entropy is TΔSTotal = 1.4 kcal⋅mol(-1) per residue at 300 K with only 20% from the loss of side-chain entropy. Our analysis exhibits mixed agreement with prior studies because of the use of more accurate ensembles and contributions from correlated motions. Buried side chains lose only a factor of 1.4 in the number of conformations available per rotamer upon folding (ΩU/ΩN). The entropy loss for helical and sheet residues differs due to the smaller motions of helical residues (TΔShelix-sheet = 0.5 kcal⋅mol(-1)), a property not fully reflected in the amide N-H and carbonyl C=O bond NMR order parameters. The results have implications for the thermodynamics of folding and binding, including estimates of solvent ordering and microscopic entropies obtained from NMR.

Physical origins of the high structural stability of CLN025 with only ten residues

CLN025, a peptide with only 10 residues, folds into a specific β-hairpin structure (this is referred to as "native structure"). Here we investigate the stabilization mechanism for CLN025 using our free-energy function F. F comprises two components, the hydration entropy and the component related to the energetic dehydration effect. The former component is calculated using the hybrid of the angle-dependent integral equation theory (ADIET) and our recently developed morphometric approach. The ADIET is a statistical-mechanical theory applied to a molecular model for water. The latter component is calculated in a simple but judicious manner accounting for physically the most important factors: the break of polypeptide-water hydrogen bonds and formation of polypeptide intramolecular hydrogen bonds upon structural change to a more compact one. We consider the native structure, compact nonnative structures newly generated, and a set of random coils mimicking the unfolded state. F and its components are calculated for all the structures considered. The loss of the polypeptide conformational entropy upon structural transition from the unfolded state to a compact structure is also estimated using a simple but physically reasonable manner. We find that the key factor is the water-entropy gain upon folding originating primarily from an increase in the total volume available to the translational displacement of water molecules in the system, which is followed by the reduction of water crowding. The amino-acid sequence of CLN025 enables it not only to closely pack the backbone and side chains including those with large aromatic groups but also to assure the intramolecular hydrogen bonding upon burial of a donor and an acceptor when the backbone forms the native structure. The assurance leads to essentially no enthalpy increase upon folding. The close packing brings a water-entropy gain which is large enough to surpass the conformational-entropy loss. By contrast, it is not possible for the design template of CLN025, GPM12, to realize the same type of structure formation. There are significantly many compact structures which are equally stable in terms of F, and due to the conformational-entropy effect, the unfolded state is favorably stabilized.

Compositional redistribution and dynamic heterogeneity in mixed lipid membrane induced by polyelectrolyte adsorption: effects of chain rigidity.

Monte Carlo simulation is employed to investigate the interaction between a polyelectrolyte and a fluid mixed membrane containing neutral (phosphatidyl-choline, PC), monovalent anionic (phosphatidylserine, PS), and multivalent anionic (phosphatidylinositol, PIP2) lipids. The effects of the intrinsic polyelectrolyte rigidity and solution ionic strength on the lateral rearrangement and dynamics of different anionic lipid species are systematically studied. Our results show that, the increase of polyelectrolyte chain rigidity reduces the loss of polyelectrolyte conformational entropy and the energy gains in electrostatic interaction, but raises the demixing entropy loss of the segregated anionic lipids. Therefore, the polyelectrolyte/membrane adsorption strength exhibits a non-monotonic dependence on the polyelectrolyte rigid parameter k ang, and there exists a certain optimal k ang for which the adsorption strength is maximal. Because the less loss of chain conformational entropy dominates the increase of the demixing entropy loss of the segregated anionic lipids and the decreases of the electrostatic energy gains, the semiflexible polyelectrolyte adsorbs onto the membrane more firmly than the flexible one. Whereas, for the adsorption of rigid polyelectrolyte, larger anionic lipid demixing entropy loss and less energy gain in the electrostatic interaction dominate over the decrease of the polyelectrolyte conformation entropy loss, leading to the desorption of the chain from the membrane. By decreasing the ionic concentration of the salt solution, the certain optimal k ang shifts to larger values. The cooperative effects of the adsorbing polyelectrolyte beads determine the concentration gradients and hierarchical mobility of the bound anionic lipids, as well as the polyelectrolyte dynamics.