The dynamics of protein hydration water: a quantitative comparison of molecular dynamics simulations and neutron-scattering experiments

We present results from an extensive molecular dynamics simulation study of water hydrating the protein Ribonuclease A, at a series of temperatures in cluster, crystal, and powder environments. The dynamics of protein hydration water appear to be very similar in crystal and powder environments at moderate to high hydration levels. Thus, we contend that experiments performed on powder samples are appropriate for discussing hydration water dynamics in native protein environments. Our analysis reveals that simulations performed on cluster models consisting of proteins surrounded by a finite water shell with free boundaries are not appropriate for the study of the solvent dynamics. Detailed comparison to available x-ray diffraction and inelastic neutron-scattering data shows that current generation force fields are capable of accurately reproducing the structural and dynamical observables. On the time scale of tens of picoseconds, at room temperature and high hydration, significant water translational diffusion and rotational motion occur. At low hydration, the water molecules are translationally confined but display appreciable rotational motion. Below the protein dynamical transition temperature, both translational and rotational motions of the water molecules are essentially arrested. Taken together, these results suggest that water translational motion is necessary for the structural relaxation that permits anharmonic and diffusive motions in proteins. Furthermore, it appears that the exchange of protein-water hydrogen bonds by water rotational/librational motion is not sufficient to permit protein structural relaxation. Rather, the complete exchange of protein-bound water molecules by translational displacement seems to be required.     

Accurate detection of protein:ligand binding sites using molecular dynamics simulations

Accurate prediction of location of cavities and surface grooves in proteins is important, as these are potential sites for ligand binding. Several currently available programs for cavity detection are unable to detect cavities near the surface or surface grooves. In the present study, an optimized molecular dynamics based procedure is described for detection and quantification of interior cavities as well as surface pockets. This is based on the observation that the mobility of water in such pockets is significantly lower than that of bulk water. The algorithm efficiently detects surface grooves that are sites of protein-ligand and protein-protein interaction. The algorithm was also used to substantially improve the performance of an automated docking procedure for docking monomers of nonobligate protein-protein complexes. In addition, it was applied to predict key residues involved in the binding of the E. coli toxin CcdB with its inhibitor. Predictions were subsequently validated by mutagenesis experiments.     

Analysis of correlated motion in antibody combining sites from molecular dynamics simulations

The crystal structures of the NC6.8-antisweet taste ligand complex and the uncomplexed antibody structures display significant differences in the conformations of residues in the combining site. A molecular dynamics method was employed to understand the flexibility and correlated motion of key combining site residues in the uncomplexed antibody. The simulations reveal that residues that show conformational differences between the complex and uncomplexed structures display strong dynamical correlations. Extensive analysis of the dynamics trajectory using time correlation methods is presented.     

Mechanical features of various silkworm crystalline considering hydration effect via molecular dynamics simulations

Silk materials are receiving significant attention as base materials for various functional nanomaterials and nanodevices, due to its exceptionally high mechanical properties, biocompatibility, and degradable characteristics. Although crystalline silk regions are composed of various repetitive motifs with differing amino acid sequences, how the effect of humidity works differently on each of the motifs and their structural characteristics remains unclear. We report molecular dynamics (MD) simulations on various silkworm fibroins composed of major motifs (i.e. (GAGAGS)_n, (GAGAGA)_n, and (GAGAGY)_n) at varying degrees of hydration, and reveal how each major motifs of silk fibroins change at each degrees of hydration using MD simulations and their structural properties in mechanical perspective via steered molecular dynamics simulations. Our results explain what effects humidity can have on nanoscale materials and devices consisting of crystalline silk materials.     

Structure of hydration water in proteins: a comparison of molecular dynamics simulations and database analysis

Hydration layer water molecules play important structural and functional roles in proteins. Despite being a critical component in biomolecular systems, characterizing the properties of hydration water poses a challenge for both experiments and simulations. In this context we investigate the local structure of hydration water molecules as a function of the distance from the protein and water molecules respectively in 188 high resolution protein structures and compare it with those obtained from molecular dynamics simulations. Tetrahedral order parameter of water in proteins calculated from previous and present simulation studies show that the potential of bulk water overestimates the average tetrahedral order parameter compared to those calculated from crystal structures. Hydration waters are found to be more ordered at a distance between the first and second solvation shell from the protein surface. The values of the order parameter decrease sharply when the water molecules are located very near or far away from the protein surface. At small water-water distance, the values of order parameter of water are very low. The average order parameter records a maximum value at a distance equivalent to the first solvation layer with respect to the water-water radial distribution and asymptotically approaches a constant value at large distances. Results from present analysis will help to get a better insight into structure of hydration water around proteins. The analysis will also help to improve the accuracy of water models on the protein surface.     

Computer simulations of protein folding by targeted molecular dynamics

We have performed 128 folding and 45 unfolding molecular dynamics runs of chymotrypsin inhibitor 2 (CI2) with an implicit solvation model for a total simulation time of 0.4 microseconds. Folding requires that the three-dimensional structure of the native state is known. It was simulated at 300 K by supplementing the force field with a harmonic restraint which acts on the root-mean-square deviation and allows to decrease the distance to the target conformation. High temperature and/or the harmonic restraint were used to induce unfolding. Of the 62 folding simulations started from random conformations, 31 reached the native structure, while the success rate was 83% for the 66 trajectories which began from conformations unfolded by high-temperature dynamics. A funnel-like energy landscape is observed for unfolding at 475 K, while the unfolding runs at 300 K and 375 K as well as most of the folding trajectories have an almost flat energy landscape for conformations with less than about 50% of native contacts formed. The sequence of events, i.e., secondary and tertiary structure formation, is similar in all folding and unfolding simulations, despite the diversity of the pathways. Previous unfolding simulations of CI2 performed with different force fields showed a similar sequence of events. These results suggest that the topology of the native state plays an important role in the folding process.     

Insights into structure, dynamics and hydration of locked nucleic acid (LNA) strand-based duplexes from molecular dynamics simulations

Locked nucleic acid (LNA) is a chemically modified nucleic acid with its sugar ring locked in an RNA-like (C3′-endo) conformation. LNAs show extraordinary thermal stabilities when hybridized with DNA, RNA or LNA itself. We performed molecular dynamics simulations on five isosequential duplexes (LNA–DNA, LNA–LNA, LNA–RNA, RNA–DNA and RNA–RNA) in order to characterize their structure, dynamics and hydration. Structurally, the LNA–DNA and LNA–RNA duplexes are found to be similar to regular RNA–DNA and RNA–RNA duplexes, whereas the LNA–LNA duplex is found to have its helix partly unwound and does not resemble RNA–RNA duplex in a number of properties. Duplexes with an LNA strand have on average longer interstrand phosphate distances compared to RNA–DNA and RNA–RNA duplexes. Furthermore, intrastrand phosphate distances in LNA strands are found to be shorter than in DNA and slightly shorter than in RNA. In case of induced sugar puckering, LNA is found to tune the sugar puckers in partner DNA strand toward C3′-endo conformations more efficiently than RNA. The LNA–LNA duplex has lesser backbone flexibility compared to the RNA–RNA duplex. Finally, LNA is less hydrated compared to DNA or RNA but is found to have a well-organized water structure.     

A Connected-cluster of hydration around myoglobin: Correlation between molecular dynamics simulations and experiment

An analysis of a molecular dynamics simulation of metmyoglobin in an explicit solvent environment of 3,128 water molecules has been performed. Both statics and dynamics of the protein-solvent interface are addressed in a comparison with experiment. Three-dimensional density distributions, temperature factors, and occupancy weights are computed for the solvent by using the trajectory coordinates. Analysis of the hydration leads to the localization of more than 500 hydration sites distributed into multiple layers of solvation located between 2.6 and 6.8 Å from the atomic protein surface. After locating the local solvent density maxima or hydration sites we conclude that water molecules of hydration positions and hydration sites are distinct concepts. Both global and detailed properties of the hydration cluster around myoglobin are compared with recent neutron and X-ray data on myoglobin. Questions arising from differences between X-ray and neutron data concerning the locations of the protein-bound water are investigated. Analysis of water site differences found from X-ray and neutron experiments compared with our simulation shows that the simulation gives a way to unify the hydration picture given by the two experiments. © 1994 John Wiley & Sons, Inc.     

Electrophoresis in protein crystal: nonequilibrium molecular dynamics simulations

Electrophoresis of a mixture of NaCl and CaCl₂ in a lysozyme crystal is investigated using nonequilibrium molecular dynamics (MD) simulations. Upon exposure to an electric field, the stability of lysozyme is found to decrease slightly. This finding is demonstrated by increases in the root mean-square deviations of the heavy atoms of lysozyme, in the solvent-accessible surface area of hydrophobic residues, and in the number of hydrogen bonds between lysozyme and water. The solvent-accessible surface area of hydrophilic residues changes marginally, and the number of hydrogen bonds between lysozyme molecules decreases. Water molecules tend to align preferentially parallel to the electric field, and the dipole moment along the pore axis increases linearly with increasing field strength. Two pronounced layered structures are observed for Na⁺ and Ca²⁺ in the vicinity of protein surface, but only one enriched layer is observed for Cl⁻. The number distributions of all ions are nearly independent of the electric field. The water coordination numbers of all ions are smaller in the crystal than in aqueous bulk solution; however, the reverse is found for the Cl⁻ coordination numbers of cations. Both the water and the Cl⁻ coordination numbers are insensitive to the electric field. Ion diffusivities in the crystal are ∼2 orders of magnitude smaller than those in aqueous bulk solution. The drift velocities of ions increase proportionally to the electric field, particularly at high strengths, and depend on ionic charge and coordination with oppositely charged ions. Electrical current exhibits a linear relationship with the field strength. The zero-field electrical conductivity is estimated to be 0.56 S/m, which is very close to 0.61 S/m as predicted by the Nernst-Einstein equation.     

Predicting thymine dimerization yields from molecular dynamics simulations

It was recently shown that thymine dimers in the all-thymine oligonucleotide (dT)₁₈ are fully formed in <1 ps after ultraviolet excitation. The speed and low quantum yield of this reaction suggest that only a small fraction of the conformers of this structurally disordered oligonucleotide are in a position to react at the instant of photon absorption. In this work, we explore the hypothesis that conventional molecular dynamics simulations can be used to predict the yield of cyclobutane pyrimidine dimers in DNA. Conformations obtained from simulations of thymidylyl-(3′-5′)-thymidine in various cosolvents were classified as dimerizable or nondimerizable depending on the distance between the C5-C6 double bonds of the adjacent thymine bases and the torsion angle between them. The quantum yield of cyclobutane pyrimidine dimer formation was calculated as the number of dimerizable conformations divided by the total number of conformations. The experimental quantum yields measured in the different solvents were satisfactorily reproduced using physically reasonable values for the two parameters. The mean dimerizable structure computed by averaging all of the dimerizable cis-syn conformations is structurally similar to the actual cis-syn dimer. Compared to the canonical B-form TT step, the most important structural property of a dimerizable conformation is its reduced helical twist angle of 22°.     

Protein flexibility using constraints from molecular dynamics simulations

Proteins are held together in the native state by hydrophobic interactions, hydrogen bonds and interactions with the surrounding water, whose strength as well as spatial and temporal distribution affects protein flexibility and hence function. We study these effects using 10 ns molecular dynamics simulations of pure water and of two proteins, the glutamate receptor ligand binding domain and barnase. We find that most of the noncovalent interactions flicker on and off over typically nanoseconds, and so we can obtain good statistics from the molecular dynamics simulations. Based on this information, a topological network of rigid bonds corresponding to a protein structure with covalent and noncovalent bonds is constructed, with account being taken of the influence of the flickering hydrogen bonds. We define the duty cycle for the noncovalent interactions as the percentage of time a given interaction is present, which we use as an input to investigate flexibility/rigidity patterns, in the algorithm FIRST which constructs and analyses topological networks.     

Interdomain dynamics and ligand binding: molecular dynamics simulations of glutamine binding protein

Periplasmic binding proteins from Gram-negative bacteria possess a common architecture, comprised of two domains linked by a hinge region, a fold which they share with the neurotransmitter-binding domains of ionotropic glutamate receptors (GluRs). Glutamine-binding protein (GlnBP) is one such protein, whose crystal structure has been solved in both open and closed forms. Multi-nanosecond molecular dynamics simulations have been used to explore motions about the hinge region and how they are altered by ligand binding. Glutamine binding is seen to significantly reduce inter-domain motions about the hinge region. Essential dynamics analysis of inter-domain motion revealed the presence of both hinge-bending and twisting motions, as has been reported for a related sugar-binding protein. Significantly, the influence of the ligand on GlnBP dynamics is similar to that previously observed in simulations of rat glutamate receptor (GluR2) ligand-binding domain. The essential dynamics analysis of GlnBP also revealed a third class of motion which suggests a mechanism for signal transmission in GluRs.     

Flexibility in a drug transport accessory protein: molecular dynamics simulations of MexA

Drug resistance in gram-negative bacteria may be conferred via efflux through a tripartite complex of an inner membrane pump, an outer membrane pore, and a periplasmic adaptor protein. These are AcrB, TolC, and AcrA, respectively, in Escherichia coli. In Pseudomonas aerugonisa, their homologs are MexB, OprM, and MexA. Defining the interdomain dynamics of the adaptor protein is essential to understanding the mechanism of complex formation. Extended (25 ns) molecular dynamics simulations of MexA have been performed to determine such interdomain dynamics. Analysis of conformational drift demonstrates substantial motions of the three domains of MexA relative to one another. Principal components analysis reveals a hinge-bending motion and rotation of the alpha-helical hairpin relative to the other domains to be the two dominant motions. These two motions provide an element of considerable flexibility which is likely to be exploited in the adaptor function of MexA.     

Anisotropic atomic motions in high-resolution protein crystallography molecular dynamics simulations

Molecular dynamics (MD) simulations using empirical force fields are popular for the study of proteins. In this work, we compare anisotropic atomic fluctuations in nanosecond-timescale MD simulations with those observed in an ultra-high-resolution crystal structure of crambin. In order to make our comparisons, we have developed a compact graphical technique for assessing agreement between spatial atomic distributions determined by MD simulations and observed anisotropic temperature factors.     

Mutations in alpha-helical solvent-exposed sites of eglin c have long-range effects: evidence from molecular dynamics simulations

Eglin c is a small protease inhibitor whose structural and thermodynamic properties have been well studied. Previous thermodynamic measurements on mutants at solvent-accessible positions in the protein's helix have shown the unexpected result that the data could be best fit by the inclusion of residue- and position-specific parameters to the model. To explore the origins of this surprising result, long molecular dynamics simulations in explicit solvent have been performed. These simulations indicate specific long-range interactions between the solvent-exposed residues in the eglin c alpha-helix and binding loop, an unexpected observation for such a small protein. The residues involved in the interaction are on opposite sides of the protein, about 25 A apart. Simulations of alanine substitutions at the solvent-exposed helix positions, arginine 22, glutamic acid 23, threonine 26, and leucine 27, show both small and large perturbations of eglin c dynamics. Two mutations exhibit large impacts on the long-range helix-loop interactions. Previous stability measurements (Yi et al., Biochemistry 2003;42:7594-7603) had indicated that an alanine substitution at position 27 was less stabilizing than at other solvent-exposed positions in the helix. The L27A mutation effects observed in these simulations suggest that the position-dependent loss of stability measured in wet bench experiments is derived from changes in dynamics that involve long-range interactions; thus, these simulations support the hypothesis that solvent-exposed positions in helices are not always equivalent.     

Efficient incorporation of protein flexibility and dynamics into molecular docking simulations

Flexibility and dynamics are protein characteristics that are essential for the process of molecular recognition. Conformational changes in the protein that are coupled to ligand binding are described by the biophysical models of induced fit and conformational selection. Different concepts that incorporate protein flexibility into protein-ligand docking within the context of these two models are reviewed. Several computational studies that discuss the validity and possible limitations of such approaches will be presented. Finally, different approaches that incorporate protein dynamics, e.g., configurational entropy, and solvation effects into docking will be highlighted.     

Misfolding pathways of the prion protein probed by molecular dynamics simulations

Although the cellular monomeric form of the benign prion protein is now well characterized, a model for the monomer of the misfolded conformation (PrP(Sc)) remains elusive. PrP(Sc) quickly aggregates into highly insoluble fibrils making experimental structural characterization very difficult. The tendency to aggregation of PrP(Sc) in aqueous solution implies that the monomer fold must be hydrophobic. Here, by using molecular dynamics simulations, we have studied the cellular mouse prion protein and its D178N pathogenic mutant immersed in a hydrophobic environment (solution of CCl4), to reveal conformational changes and/or local structural weaknesses of the prion protein fold in unfavorable structural and thermodynamic conditions. Simulations in water have been also performed. Although observing in general a rather limited conformation activity in the nanosecond timescale, we have detected a significant weakening of the antiparallel beta-sheet of the D178N mutant in CCl4 and to a less extent in water. No weakening is observed for the native prion protein. The increase of beta-structure in the monomer, recently claimed as evidence for misfolding to PrP(Sc), has been also observed in this study irrespective of the thermodynamic or structural conditions, showing that this behavior is very likely an intrinsic characteristic of the prion protein fold.