Parallel tempering molecular dynamics folding simulation of a signal peptide in explicit water

Parallel temperature molecular dynamics simulations are used to explore the folding of a signal peptide, a short but functionally independent domain at the N-terminus of proteins. The peptide has been analyzed previously by NMR, and thus a solid reference state is provided with the experimental structure. Particular attention is paid to the role of water considered in full atomic detail. Different partial aspects in the folding process are quantified. The major group of obtained structures matches the NMR structure very closely. An important biological consequence is that in vivo folding of signal peptides seems to be possible within aqueous environments.     

Simulations of apo and holo-fatty acid binding protein: structure and dynamics of protein, ligand and internal water.

Two molecular dynamics simulations of 5 ns each have been carried out for rat intestinal fatty acid binding protein, in apo-form and with bound palmitate. The fatty acid and a number of water molecules are encapsulated in a large interior cavity of the barrel-shaped protein. The simulations are compared to experimental data and analyzed in terms of root mean square deviations, atomic B-factors, secondary structure elements, hydrogen bond patterns, and distance constraints derived from nuclear Overhauser experiments. Excellent agreement is found between simulated and experimental solution structures of holo-FABP, but a number of differences are observed for the apo-form. The ligand in holo-FABP shows considerable displacement after about 1.5 ns and displays significant configurational entropy. A novel computational approach has been employed to identify internal water and to capture exchange pathways. Orifices in the portal and gap regions of the protein, discussed in the experimental literature, have been confirmed as major openings for solvent exchange between the internal cavity and bulk water. A third opening on the opposite side of the barrel experiences significant exchange but it does not provide a pathway for further passage to the central cavity. Internal water is characterized in terms of density distributions, interaction energies, mobility, protein contact times, and water molecule coordination. A number of differences are observed between the apo and holo-forms and related to differences in the protein structure. Solvent inside apo-FABP, for example, shows characteristics of a water droplet, while solvent in holo-FABP benefits from interactions with the ligand headgroup and slightly stronger interactions with protein residues.     

The influence of molecular variation on protein interactions

We investigated the influence of solvation forces on protein-protein interactions for two forms of lysozyme: hen egg white (HEWL) and turkey egg white (TEWL). Turkey egg white has more surface exposed hydrophobic residues than HEWL and the protein-protein interactions of TEWL are shown to be more attractive than those of HEWL, for the conditions studied. The importance of including a solvation term in the potential of mean force model, to account for molecular variation in protein surface characteristics, is highlighted. We also show that the magnitude of this solvation term can be estimated using readily available data.     

Non‐interacting surface solvation and dynamics in protein–protein interactions

Protein–protein interactions control a plethora of cellular processes, including cell proliferation, differentiation, apoptosis, and signal transduction. Understanding how and why proteins interact will inevitably lead to novel structure‐based drug design methods, as well as design of de novo binders with preferred interaction properties. At a structural and molecular level, interface and rim regions are not enough to fully account for the energetics of protein–protein binding, even for simple lock‐and‐key rigid binders. As we have recently shown, properties of the global surface might also play a role in protein–protein interactions. Here, we report on molecular dynamics simulations performed to understand solvent effects on protein–protein surfaces. We compare properties of the interface, rim, and non‐interacting surface regions for five different complexes and their free components. Interface and rim residues become, as expected, less mobile upon complexation. However, non‐interacting surface appears more flexible in the complex. Fluctuations of polar residues are always lower compared with charged ones, independent of the protein state. Further, stable water molecules are often observed around polar residues, in contrast to charged ones. Our analysis reveals that (a) upon complexation, the non‐interacting surface can have a direct entropic compensation for the lower interface and rim entropy and (b) the mobility of the first hydration layer, which is linked to the stability of the protein–protein complex, is influenced by the local chemical properties of the surface. These findings corroborate previous hypotheses on the role of the hydration layer in shielding protein–protein complexes from unintended protein–protein interactions. Proteins 2015; 83:445–458. © 2014 Wiley Periodicals, Inc.     

Influence of divalent magnesium ion on DNA: molecular dynamics simulation studies

A large amount of experimental evidence is available on the effect of magnesium ions on the structure and stability of DNA double helix. Less is known, however, on how these ions affect the stability and dynamics of the molecule. The static time average pictures from X-ray structures or the quantum chemical energy minimized structures lack understanding of the dynamic DNA–ion interaction. The present work addresses these questions by molecular dynamics simulation studies on two DNA duplexes and their interaction with magnesium ions. Results show typical B-DNA character with occasional excursions to deviated states. We detected expected stability of the duplexes in terms of backbone conformations and base pair parameter by the CHARMM-27 force field. Ion environment analysis shows that Mg²⁺ retains the coordination sphere throughout the simulation with a preference for major groove over minor. An extensive analysis of the influence of the Mg²⁺ ion shows no evidence of the popular predictions of groove width narrowing by dipositive metal ion. The major groove atoms show higher occupancy and residence time compared to minor groove for magnesium, where no such distinction is found for the charge neutralizing Na⁺ ions. The determining factor of Mg²⁺ ion’s choice in DNA binding site evolves as the steric hindrance faced by the bulky hexahydrated cation where wider major groove gets the preference. We have shown that in case of binding of Mg²⁺ to DNA non electrostatic contributions play a major role.

An animated Interactive 3D Complement (I3DC) is available in Proteopedia at http://proteopedia.org/w/Journal:JBSD:5     

Ions and counterions in a biological channel: a molecular dynamics simulation of OmpF porin from Escherichia coli in an explicit membrane with 1 M KCl aqueous salt solution

A 5 ns all-atom molecular dynamics trajectory of Escherichia coli OmpF porin embedded in an explicit dimyristoyl-phosphatidylcholine (DMPC) bilayer bathed by a 1 M [KCl] aqueous salt solution is generated to explore the microscopic details of the mechanism of ion permeation. The atomic model includes the OmpF trimer, 124 DMPC, 13470 water molecules as well as 231 K+ and 201 Cl-, for a total of 70,693 atoms. The structural and dynamical results are in excellent agreement with the X-ray data. The global root-mean-square deviation of the backbone atoms relative to the X-ray structure is 1.4 A. A cluster of three fully charged arginine (Arg42, Arg82, and Arg132) facing two acidic residues (Asp113 and Glu117) on L3 in the narrowest part of the aqueous pore is observed to be very stable in the crystallographic conformation. In this region of the pore, the water molecules are markedly oriented perpendicular to the channel axis due to the strong transversal electrostatic field arising from those residues. On average the size of the pore is smaller during the simulation than in the X-ray structure, undergoing small fluctuations. No large movements of loop L3 leading to a gating of the pore are observed. Remarkably, it is observed that K+ and Cl- follow two well-separated average pathways spanning over nearly 40 A along the axis of the pore. In the center of the monomer, the two screw-like pathways have a left-handed twist, undergoing a counter-clockwise rotation of 180 degrees from the extracellular vestibule to the pore periplasmic side. In the pore, the dynamical diffusion constants of the ions are reduced by about 50% relative to their value in bulk solvent. Analysis of ion solvation across the channel reveals that the contributions from the water and the protein are complementary, keeping the total solvation number of both ions nearly constant. Unsurprisingly, K+ have a higher propensity to occupy the aqueous pore than Cl-, consistent with the cation selectivity of the channel. However, further analysis suggests that ion-ion pairs play an important role. In particular, it is observed that the passage of Cl- occurs only in the presence of K+ counterions, and isolated K+ can move through the channel and permeate on their own. The presence of K+ in the pore screens the negative electrostatic potential arising from OmpF to help the translocation of Cl- by formation of ion pairs.     

Studying pressure denaturation of a protein by molecular dynamics simulations

Many globular proteins unfold when subjected to several kilobars of hydrostatic pressure. This “unfolding‐up‐on‐squeezing” is counter‐intuitive in that one expects mechanical compression of proteins with increasing pressure. Molecular simulations have the potential to provide fundamental understanding of pressure effects on proteins. However, the slow kinetics of unfolding, especially at high pressures, eliminates the possibility of its direct observation by molecular dynamics (MD) simulations. Motivated by experimental results—that pressure denatured states are water‐swollen, and theoretical results—that water transfer into hydrophobic contacts becomes favorable with increasing pressure, we employ a water insertion method to generate unfolded states of the protein Staphylococcal Nuclease (Snase). Structural characteristics of these unfolded states—their water‐swollen nature, retention of secondary structure, and overall compactness—mimic those observed in experiments. Using conformations of folded and unfolded states, we calculate their partial molar volumes in MD simulations and estimate the pressure‐dependent free energy of unfolding. The volume of unfolding of Snase is negative (approximately ?60 mL/mol at 1 bar) and is relatively insensitive to pressure, leading to its unfolding in the pressure range of 1500–2000 bars. Interestingly, once the protein is sufficiently water swollen, the partial molar volume of the protein appears to be insensitive to further conformational expansion or unfolding. Specifically, water‐swollen structures with relatively low radii of gyration have partial molar volume that are similar to that of significantly more unfolded states. We find that the compressibility change on unfolding is negligible, consistent with experiments. We also analyze hydration shell fluctuations to comment on the hydration contributions to protein compressibility. Our study demonstrates the utility of molecular simulations in estimating volumetric properties and pressure stability of proteins, and can be potentially extended for applications to protein complexes and assemblies. Proteins 2010. © 2009 Wiley‐Liss, Inc.     

Exploring the influence of hyperthermophilic protein Ssh10b on the stability and conformation of RNA by molecular dynamics simulation

The hyperthermophilic Ssh10b from Sulfolobus shibatae is a member of the Sac10b family, which binds RNA in vivo as a physiological substrate, and it has been postulated to play a key role in chromosomal organization in Archaea. Even though the crystal structure of Ssh10b‐RNA was resolved successively by X‐ray diffraction (Protein Data Bank [PDB] code: 3WBM), the detailed dynamic characteristics of Ssh10b‐RNA are still unclear. In this study, molecular dynamics (MDs) simulations at 6 temperatures (300, 350, 375, 400, 450, and 500 K) and molecular mechanics Generalized‐Born surface area (MM‐GB/SA) free energy calculations were performed to investigate the mechanism of how Ssh10b protects and stabilizes RNA. The simulation results indicate that RNA is stabilized by Ssh10b when the temperature rises up to 375 K. RNA is found to undergo conformational transition between A‐RNA and A′‐RNA when Ssh10b binds to RNA at 3 different temperatures (300, 350, and 375 K). Salt bridges, hydrogen bonds and hydrophobic interactions are observed, and some residues have significant impact on the structural stability of the complex. This study increases our understanding of the dynamics and interaction mechanism of hyperthermophilic proteins and RNA at the atomic level, and offers a model for studying the structural biology of hyperthermophilic proteins and RNA.     

Standards for molecular dynamics modelling and simulation of relaxation

An attempt is made to formulate a set of requirements for simulation and modelling of relaxation in dense media. Each requirement is illustrated by examples of numerical simulation of particles with different types of interaction given by soft-sphere, Lennard–Jones, embedded atom method or Coulomb potential. The approaches developed are expected to be universal for some classes of relaxation processes in liquids, fluids, crystals and plasmas.     

Transport diffusivities of fluids in nanopores by non-equilibrium molecular dynamics simulation

We present a method to study fluid transport through nanoporous materials using highly efficient non-equilibrium molecular dynamics simulations. A steady flow is induced by applying an external field to the fluid particles within a small slab of the simulation cell. The external field generates a density gradient between both sides of the porous material, which in turn triggers a convective flux through the porous medium. The heat dissipated by the fluid flow is released by a Gaussian thermostat applied to the wall particles. This method is effective for studying diffusivities in a slit pore as well as more natural, complex wall geometries. The dependence of the diffusive flux on the external field sheds light on the transport diffusivities and allows a direct calculation of effective diffusivities. Both pore and fluid particle interactions are represented by coarse-grained molecular models in order to present a proof-of-concept and to retain computational efficiency in the simulations. The application of the method is demonstrated in two different scenarios, namely the effective mass transport through a slit pore and the calculation of the effective self-diffusion through this system. The method allows for a distinction between diffusive and convective contributions of the mass transport.     

Distal mutation modulates the heme sliding in mouse neuroglobin investigated by molecular dynamics simulation

Neuroglobin (Ngb), a hexa‐coordinated hemoprotein primarily expressed in the brain and retina, is thought to be involved in neuroprotection and signal transduction. Ngb can reversibly bind small ligands such as O₂ and CO to the heme iron by replacing the distal histidine which is bound to the iron as the endogenous ligand. In this work, molecular dynamics (MD) simulations were performed to investigate the functionally related structural properties and dynamical characteristics in carboxy mouse neuroglobin and three distal mutants including single mutants H64V, K67T and double mutant H64V/K67T. MD simulations suggest that the heme sliding motion induced by the binding of exogenous ligand is affected by the distal mutation obviously. Accompanying changes in loop flexibility and internal cavities imply the structural rearrangement of Ngb. Moreover, the solvent accessibility of heme and some crucial residues are influenced revealing an interactive network on the distal side. The work elucidates that the key residues K67 at E10 and H64 at E7 are significant in modulating the heme sliding and hence the structural and physiological function of Ngb. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

Phase-transition properties of glycerol-monopalmitate lipid bilayers investigated by molecular dynamics simulation: influence of the system size and force-field parameters

Phase-transition properties of glycerol-1-monopalmitate (GMP) bilayers are investigated using explicit-solvent molecular dynamics (MD) simulations, initiated from structures appropriate for the gel (GL) or liquid crystal (LC) phases, and carried out at different hydration levels and temperatures. Building up on a previous study and based on 600 ns simulations, the influence of the system size and of the force field on the equilibrium thermodynamic and dynamic parameters of the bilayers in the GL and LC phases, as well as on the temperature T_m and properties of the GL ? LC phase transition, are analysed. Qualitatively speaking, the results agree with the available experimental data for the area per lipid in the two phases and for the phase-transition temperatures at the three hydration levels irrespective of the selected model parameters. They also suggest that the total number of hydrogen bonds formed between a lipid headgroup and its environment is essentially constant, amounting to about four in both the LC and the GL phases. Quantitatively speaking, the dependence of T_m on the hydration level is found to be non-systematic across the different combinations of model parameters. This results in part from a sensitivity of the results on the system size and force-field parameters but also from the limited accuracy of the bracketing approach employed here to estimate T_m. Finally, a simple kinetic model is proposed to account for the timescales of the transitions. This model involves enthalpy and entropy increases of about 26 kJ mol^? 1 and 83 J mol^? 1 K^? 1 per lipid, upon going from the GL to the LC phase. The transition state is associated with activation parameters corresponding to 13% and 11%, respectively, of these values along the GL → LC transition, resulting in an activation free energy of about 0.3 kJ mol^? 1 per lipid at T_m.     

Structure and dynamics of cumene and 1,2,4-trimethylbenzene mixture in NaY zeolite: a molecular dynamics simulation study

A molecular dynamics study of 1:1 mixture of the C9 isomers, isopropyl benzene (cumene) and 1,2,4-trimethylbenzene (124TMB) (pseudocumene), in zeolite NaY is reported. Structural and dynamical properties have been computed to understand possible difficulties in the separation of these isomers. Cumene exhibits a slightly higher self-diffusivity. 124TMB encounters a larger barrier as compared with cumene at the 12-ring window during migration from one supercage to another. 124TMB has a significantly larger backscattering during rotation which may be attributed to its shape and large cross-sectional diameter as compared with cumene. Cumene has a higher rotational diffusivity. Results suggest that there is larger difference between the rotational diffusivities of the two isomers and little difference in their translational diffusivity. It may be possible to exploit this difference in separating the two isomers.     

Investigation of moisture diffusion in cross-linked epoxy moulding compound by molecular dynamics simulation

Molecular dynamics (MD) simulation was carried out to study the moisture diffusion in cross-linked epoxy resin, with the influence of temperature, water concentration and polymer conversion taken into account. The simulation results showed that the moisture diffusion coefficients increased with the increase in temperature. And generally, with the increased moisture concentration or decreased polymer conversion, the moisture diffusion coefficients reduced. However, the moisture diffusion was strongly inhibited when the number of epoxy groups in completely reacted epoxy resins was equal to the number of water molecules.     

Water dynamics in the large cavity of three lipid-binding proteins monitored by (17)O magnetic relaxation dispersion

Intracellular lipid-binding proteins contain a large binding cavity filled with water molecules. The role played by these water molecules in ligand binding is not well understood, but their energetic and dynamic properties must be important for protein function. Here, we use the magnetic relaxation dispersion (MRD) of the water 17O resonance to investigate the water molecules in the binding cavity of three different lipid-binding proteins: heart fatty acid-binding protein (H-FABP), ileal lipid-binding protein (I-LBP) and intestinal fatty acid-binding protein (I-FABP). Whereas about half of the crystallographically visible water molecules appear to be expelled by the ligand, we find that ligand binding actually increases the number of water molecules within the cavity. At 300 K, the water molecules in the cavity exchange positions on a time-scale of about 1ns and exchange with external water on longer time-scales (0.01-1 micros). Exchange of water molecules among hydration sites within the cavity should be strongly coupled to ligand motion. Whereas a recent MD simulation indicates that the structure of the cavity water resembles a bulk water droplet, the present MRD results show that its dynamics is more than two orders of magnitude slower than in the bulk. These findings may have significant implications for the strength, specificity and kinetics of lipid binding.     

Structure of the antimicrobial beta-hairpin peptide protegrin-1 in a DLPC lipid bilayer investigated by molecular dynamics simulation

All atom molecular dynamics simulations of the 18-residue beta-hairpin antimicrobial peptide protegrin-1 (PG-1, RGGRLCYCRRRFCVCVGR-NH(2)) in a fully hydrated dilauroylphosphatidylcholine (DLPC) lipid bilayer have been implemented. The goal of the reported work is to investigate the structure of the peptide in a membrane environment (previously solved only in solution [R.L. Fahrner, T. Dieckmann, S.S.L. Harwig, R.I. Lehrer, D. Eisenberg, J. Feigon, Solution structure of protegrin-1, a broad-spectrum antimicrobial peptide from porcine leukocytes. Chemistry and Biology, 3 (1996) 543-550]), and to delineate specific peptide-membrane interactions which are responsible for the peptide's membrane binding properties. A novel, previously unknown, "kick" shaped conformation of the peptide was detected, where a bend at the C-terminal beta-strand of the peptide caused the peptide backbone at residues 16-18 to extend perpendicular to the beta-hairpin plane. This bend was driven by a highly persistent hydrogen-bond between the polar peptide side-chain of TYR7 and the unshielded backbone carbonyl oxygen atom of GLY17. The H-bond formation relieves the unfavorable free energy of insertion of polar groups into the hydrophobic membrane core. PG-1 was anchored to the membrane by strong electrostatic binding of the protonated N-terminus of the peptide to the lipid head group phosphate anions. The orientation of the peptide in the membrane, and its influence on bilayer structural and dynamic properties are in excellent agreement with solid state NMR measurements [S. Yamaguchi, T. Hong, A. Waring, R.I. Lehrer, M. Hong, Solid-State NMR Investigations of Peptide-Lipid Interaction and Orientation of a b-Sheet Antimicrobial Peptide, Protegrin, Biochemistry, 41 (2002) 9852-9862]. Importantly, two simulations which started from different initial orientations of the peptide converged to the same final equilibrium orientation of the peptide relative to the bilayer. The kick-shaped conformation was observed only in one of the two simulations.     

Topological and dynamical properties of Azurin anchored to a gold substrate as investigated by molecular dynamics simulation

A classical molecular dynamics study of the electron transfer protein azurin, covalently bound to a gold substrate through its native disulphide group, is carried out at full hydration. With the aim of investigating the effects on the protein structure and dynamics as induced by the presence of an electric field, simulations are performed on neutral, positively and negatively charged substrates. A number of parameters, such as the average structure, the root mean square deviations and fluctuations, the intraprotein hydrogen bonds and solvent accessible surface of the protein, are monitored during 10 ns of run. The orientation, the height and the lateral size of the protein, with respect to the substrate are evaluated and compared with the experimental data obtained by scanning probe nanoscopies. The electron transfer properties between the copper redox center and the disulphide bridge bound to the substrate are investigated and briefly discussed.     

Effect of hexafluoroisopropanol alcohol on the structure of melittin: a molecular dynamics simulation study

The molecular mechanism by which HFIP stabilizes the alpha-helical structure of peptides is not well understood. In the present study, we use melittin as a model to gain insight into the details of the atomistic interactions of HFIP with the peptide. We have performed extensive comparative molecular dynamics simulations (up to 100 nsec) in the absence and in the presence of HFIP. In agreement with recent NMR experiments, the simulations show rapid loss of tertiary structure in water at pH 2 but much higher helicity in 35% HFIP. The MD simulations also indicate that melittin adopts a highly dynamic global structure in 35% HFIP solution with two alpha-helical segments sampling a wide range of angular orientations. The analysis of the HFIP distribution shows the tendency of HFIP to aggregate around the peptide, increasing the local cosolvent concentration to more than two times that in the bulk concentration. The correlation of local peptide structure with HFIP coating suggests that displacement of water at the peptide surface is the main contribution of HFIP in stabilizing the secondary structure of melittin. Finally, a stabilizing effect promoted by the presence of counter-ions was also observed in the simulations.     

Toward understanding the inactivation mechanism of monooxygenase P450 BM-3 by organic cosolvents: a molecular dynamics simulation study

Cytochrome P450 BM-3 from Bacillus megaterium is an extensively studied enzyme for industrial applications. A major focus of current protein engineering research is directed to improving the catalytic performance of P450 BM-3 toward nonnatural substrates of industrial importance in the presence of organic solvents or cosolvents. For the latter reason, it is important to study the effect of organic cosolvent molecules on the structure and dynamics of the enzyme, in particular, the effect of cosolvent molecules on the active site's structure and dynamics. In this paper, we have studied, using molecular dynamics (MD) simulations, the F87A mutant of P450 BM-3 in the presence of DMSO as cosolvent, to understand the role of the F87A substitution for its catalytic activity. This mutant exhibits an altered regioselectivity and substrate specificity compared with wild-type; however, it has lower tolerance toward DMSO. The simulation results offer an explanation for the DMSO sensitivity of the F87A mutant. Our simulation results show that the F87 side chain prevents the disturbance of the water molecule bound to the heme iron by DMSO molecules. The absence of the phenyl ring in F87A mutant promotes interactions of the DMSO molecule with the heme iron resulting in water displacement by DMSO at the catalytic heme center.     

The role of flexibility and hydration on the sequence-specific DNA recognition by the Tn916 integrase protein: a molecular dynamics analysis

The N-terminal domain of the Tn916 integrase protein (INT-DBD) is responsible for DNA binding in the process of strand cleavage and joining reactions required for transposition of the Tn916 conjugative transposon. Site-specific association is facilitated by numerous protein-DNA contacts from the face of a three-stranded beta-sheet inserted into the major groove. The protein undergoes a subtle conformational transition and is slightly unfolded in the protein-DNA complex. The conformation of many charged residues is poorly defined by NMR data but mutational studies have indicated that removal of polar side chains decreases binding affinity, while non-polar contacts are malleable. Based on analysis of the binding enthalpy and binding heat capacity, we have reasoned that dehydration of the protein-DNA interface is incomplete. This study presents results from a molecular dynamics investigation of the INT-DBD-DNA complex aimed at a more detailed understanding of the role of conformational dynamics and hydration in site-specific binding. Comparison of simulations (total of 13 ns) of the free protein and of the bound protein conformation (in isolation or DNA-bound) reveals intrinsic flexibility in certain parts of the molecule. Conformational adaptation linked to partial unfolding appears to be induced by protein-DNA contacts. The protein-DNA hydrogen-bonding network is highly dynamic. The simulation identifies protein-DNA interactions that are poorly resolved or only surmised from the NMR ensemble. Single water molecules and water clusters dynamically optimize the complementarity of polar interactions at the 'wet' protein-DNA interface. The simulation results are useful to establish a qualitative link between experimental data on individual residue's contribution to binding affinity and thermodynamic properties of INT-DBD alone and in complex with DNA.