The effect of cholesterol on short- and long-chain monounsaturated lipid bilayers as determined by molecular dynamics simulations and X-ray scattering

We investigate the structure of cholesterol-containing membranes composed of either short-chain (diC14:1PC) or long-chain (diC22:1PC) monounsaturated phospholipids. Bilayer structural information is derived from all-atom molecular dynamics simulations, which are validated via direct comparison to x-ray scattering experiments. We show that the addition of 40 mol % cholesterol results in a nearly identical increase in the thickness of the two different bilayers. In both cases, the chain ordering dominates over the hydrophobic matching between the length of the cholesterol molecule and the hydrocarbon thickness of the bilayer, which one would expect to cause a thinning of the diC22:1PC bilayer. For both bilayers there is substantial headgroup rearrangement for lipids directly in contact with cholesterol, supporting the so-called umbrella model. Importantly, in diC14:1PC bilayers, a dynamic network of hydrogen bonds stabilizes long-lived reorientations of some cholesterol molecules, during which they are found to lie perpendicular to the bilayer normal, deep within the bilayer’s hydrophobic core. Additionally, the simulations show that the diC14:1PC bilayer is significantly more permeable to water. These differences may be correlated with faster cholesterol flip-flop between the leaflets of short-chain lipid bilayers, resulting in an asymmetric distribution of cholesterol molecules. This asymmetry was observed experimentally in a case of unilamellar vesicles (ULVs), and reproduced through a set of novel asymmetric simulations. In contrast to ULVs, experimental data for oriented multilamellar stacks does not show the asymmetry, suggesting that it results from the curvature of the ULV bilayers.     

Multiple pH regime molecular dynamics simulation for pK calculations

Ionisation equilibria in proteins are influenced by conformational flexibility, which can in principle be accounted for by molecular dynamics simulation. One problem in this method is the bias arising from the fixed protonation state during the simulation. Its effect is mostly exhibited when the ionisation behaviour of the titratable groups is extrapolated to pH regions where the predetermined protonation state of the protein may not be statistically relevant, leading to conformational sampling that is not representative of the true state. In this work we consider a simple approach which can essentially reduce this problem. Three molecular dynamics structure sets are generated, each with a different protonation state of the protein molecule expected to be relevant at three pH regions, and pK calculations from the three sets are combined to predict pK over the entire pH range of interest. This multiple pH molecular dynamics approach was tested on the GCN4 leucine zipper, a protein for which a full data set of experimental data is available. The pK values were predicted with a mean deviation from the experimental data of 0.29 pH units, and with a precision of 0.13 pH units, evaluated on the basis of equivalent sites in the dimeric GCN4 leucine zipper.     

Physicochemical properties of blue fluorescent protein determined via molecular dynamics simulation

Blue fluorescent protein (BFP) is a mutant of green fluorescent protein (GFP), where the chromophore has been modified to shift the emitted fluorescence into the blue spectral region. In this study, MD calculations were performed with the GROMACS simulation package and AMBER force field to investigate the dependence of BFPs physicochemical properties on temperature and applied pressure. The MD approach enabled us to calculate the compressibility of protein itself, separately from the nontrivial contribution of the hydration shell, which is difficult to achieve experimentally. The computed compressibility of BFP (3.94 x10(-5) MPa(-1)) is in agreement with experimental values of globular proteins. The center-of-mass diffusion coefficient of BFP and its dependence on temperature and pressure, which plays an important role in its application as a probe for intracellular liquid viscosity measurement, was calculated and found to be in good agreement with photobleaching recovery experimental data. We have shown that decreased temperature as well as applied pressure increases the water viscosity, but the concomitant decrease of the BFP diffusion coefficient behaves differently from Stokes-Einstein formula. It is shown that the number of hydrogen bonds around the protein grows with pressure, which explains the aforementioned deviation. Pressure also reduces root mean square (RMS) fluctuations, especially those of the most flexible residues situated in the loops. The analysis of the RMS fluctuations of the backbone Calpha atoms also reveals that the most rigid part of BFP is the center of the beta-barrel, in accord with temperature B factors obtained from the Protein Data Bank.     

The open state of human topoisomerase I as probed by molecular dynamics simulation

The open state of human topoisomerase I has been probed by molecular dynamics simulation, starting from the coordinates of the closed structure of the protein complexed with DNA, after elimination of the 22-bp DNA duplex oligonucleotide. A repulsion force between the two lips of the protein has been introduced for a short time to induce destabilization of the local minimum, after which an unperturbed simulation has been carried out for 10ns. The simulation shows that the protein undergoes a large conformational change due to rearrangements in the orientation of the protein domains, which however move as a coherent unit, fully maintaining their secondary and tertiary structures. Despite movements between the domains as large as 80–90Å, the catalytic pentad remains preassembled, the largest deviation of the active site backbone atoms from the starting crystallographic structure being only 1.7Å. Electrostatic calculation of the open protein structure shows that the protein displays a vast positive region with the active site residues located nearly at its center, in a conformation perfectly suited to interact with the negatively charged supercoiled DNA substrate.     

Exploring the effect of inhibitor AKB-9778 on VE-PTP by molecular docking and molecular dynamics simulation

Diabetic macular edema, also known as diabetic eye disease, is mainly caused by the overexpression of vascular endothelial protein tyrosine phosphatase (VE-PTP) at hypoxia/ischemic. AKB-9778 is a known VE-PTP inhibitor that can effectively interact with the active site of VE-PTP to inhibit the activity of VE-PTP. However, the binding pattern of VE-PTP with AKB-9778 and the dynamic implications of AKB-9778 on VE-PTP system at the molecular level are poorly understood. Through molecular docking, it was found that the AKB-9778 was docked well in the binding pocket of VE-PTP by the interactions of hydrogen bond and Van der Waals. Furthermore, after molecular dynamic simulations on VE-PTP system and VE-PTP ^AKB-9778 system, a series of postdynamic analyses found that the flexibility and conformation of the active site undergone an obvious transition after VE-PTP binding with AKB-9778. Moreover, by constructing the RIN, it was found that the different interactions in the active site were the detailed reasons for the conformational differences between these two systems. Thus, the finding here might provide a deeper understanding of AKB-9778 as VE-PTP Inhibitor.     

Relative stability of protein structures determined by X-ray crystallography or NMR spectroscopy: a molecular dynamics simulation study

The relative stability of protein structures determined by either X-ray crystallography or nuclear magnetic resonance (NMR) spectroscopy has been investigated by using molecular dynamics simulation techniques. Published structures of 34 proteins containing between 50 and 100 residues have been evaluated. The proteins selected represent a mixture of secondary structure types including all alpha, all beta, and alpha/beta. The proteins selected do not contain cysteine-cysteine bridges. In addition, any crystallographic waters, metal ions, cofactors, or bound ligands were removed before the systems were simulated. The stability of the structures was evaluated by simulating, under identical conditions, each of the proteins for at least 5 ns in explicit solvent. It is found that not only do NMR-derived structures have, on average, higher internal strain than structures determined by X-ray crystallography but that a significant proportion of the structures are unstable and rapidly diverge in simulations.     

Yeast biodiversity and dynamics during sweet wine production as determined by molecular methods

In this study we investigated yeast biodiversity and dynamics during the production of a sweet wine obtained from dried grapes. Two wineries were selected in the Collio region and grapes, grape juices and wines during fermentations were analyzed by culture-dependent methods (plating on WLN medium) and culture-independent methods (PCR-DGGE). Moreover, the capability of the Saccharomyces cerevisiae starter cultures to take over the fermentation was assessed by RAPD-PCR. On WLN agar several species of non-Saccharomyces yeasts (Hanseniaspora, Metschnikowia, Pichia, Candida, Torulaspora and Debaryomyces), but also strains of S. cerevisiae, were isolated. After inoculation of the starter cultures, only colonies typical of S. cerevisiae were observed. Using PCR-DGGE, the great biodiversity of moulds on the grapes was underlined, both at the DNA and RNA level, while the yeast contribution started to become important only in the musts. Here, bands belonging to species of Candida zemplinina and Hanseniaspora uvarum were visible. Lastly, when the S. cerevisiae isolates were compared by RAPD-PCR, it was determined that only in one of the fermentations followed, the inoculated strain conducted the alcoholic fermentation. In the second fermentation, the starter culture was not able to promptly implant and other populations of S. cerevisiae could be isolated, most likely contributing to the final characteristics of the sweet wine produced.     

The effect of alumina nanoparticles on the thermal properties of PMMA: a molecular dynamics simulation

Metal oxides, as one of the most promising flame retardant additives, improve the fire retardant and the thermal stability properties of polymers. In the present study, molecular dynamics (MD) simulations based on the united atom model were applied to study the effect of alumina nanoparticles on the density, thermal conductivity, heat capacity, and thermal diffusivity of isotactic poly(methyl methacrylate) (is-PMMA). Thermal diffusivity of PMMA and PMMA/alumina nanocomposite were investigated through calculating thermal conductivity, density and heat capacity in the range of 300–700?K. Heat capacity can be calculated using fluctuations properties of energy. Thermal conductivity was calculated through the nonequilibrium molecular dynamics (NEMD) simulation by Fourier’s law approach. Our results show that the addition of alumina nanoparticles decreases the heat capacity and increases the glass transition temperature (T_g), thermal conductivity and thermal diffusivity of the PMMA. Therefore, the addition of alumina nanoparticles to PMMA improves the fire retardancy of the polymer. In addition, we illustrate the links between the intermolecular and bulk properties of PMMA in the presence of the alumina nanoparticles.     

The mechanism for the complexation and dissociation between siRNA and PMAL: a molecular dynamics simulation study based on a coarse-grained model

Abstract

The capacity of silencing genes makes small interfering RNA (siRNA) becomes potential candidates for curing many fatal diseases. Due to the low stability and delivery efficiency of siRNA, the design of amphiphilic carrier for siRNA delivery is vital for the practical gene therapy. In the present work, we explored how the complexation and dissociation of siRNA with poly (maleic anhydride-alt-1-decene) substituted with 3-(dimethylamino) propylamine (PMAL), which is a recent synthesised amphiphilic polymer and can be used in delivery of siRNAs and proteins, using traditional molecular dynamics simulations, together with steered molecular dynamics simulations. It was shown that the complexation of siRNA with PMALs can spontaneously occur, no matter what unit number of PMAL is. PMALs of different unit numbers form micelle-like structures and interact with siRNA surface. With the increase of unit number, PMAL becomes more flexible and interacts with siRNA from attachment to entanglement. The dissociation of PMAL from siRNA is an energy-consuming process. The free energy difference increases with the unit number of PMAL. The free energy for dissociation involves both the stretch of PMAL and the separation of PMAL from siRNA. Therefore, an optimal unit number of PMAL is critical for the delivery efficiency of siRNA when PMAL is used as carrier. In present work, when the radius of gyration of PMAL approaches to that of siRNA, PMAL gives a favoured both complexation and dissociation between siRNA and PMAL. Finally, we propose the mechanism of complexation and dissociation of PMAL with siRNA. The above simulation established a molecular insight of the interaction between siRNA and PMAL and was helpful for the design and applications of new PMAL-based polymers as siRNA delivery carriers.     

Theoretical considerations on the role of membrane potential in the regulation of endosomal pH.

Na+,K(+)-ATPase has been observed to partially inhibit acidification of early endosomes by increasing membrane potential, whereas chloride channels have been observed to enhance acidification in endosomes and lysosomes. However, little theoretical analysis of the ways in which different pumps and channels may interact has been carried out. We therefore developed quantitative models of endosomal pH regulation based on thermodynamic considerations. We conclude that 1) both size and shape of endosomes will influence steady-state endosomal pH whenever membrane potential due to the pH gradient limits proton pumping, 2) steady-state pH values similar to those observed in early endosomes of living cells can occur in endosomes containing just H(+)-ATPases and Na+,K(+)-ATPases when low endosomal buffering capacities are present, and 3) inclusion of active chloride channels results in predicted pH values well below those observed in vivo. The results support the separation of endocytic compartments into two classes, those (such as early endosomes) whose acidification is limited by attainment of a certain membrane potential, and those (such as lysosomes) whose acidification is limited by the attainment of a certain pH. The theoretical framework and conclusions described are potentially applicable to other membrane-enclosed compartments that are acidified, such as elements of the Golgi apparatus.     

Molecular dynamics simulation and steered molecular dynamics simulation on irisin dimers

Irisin is found closely associated with promoting the browning of beige fat cells in white adipose tissue. The crystal structure reveals that irisin forms a continuous inter-subunit β-sheet dimer. Here, molecular dynamics (MD) simulation and steered molecular dynamics (SMD) simulation were performed to investigate the dissociation process and the intricate interactions between the two irisin monomers. In the process of MD, the interactions between the monomers were roughly analyzed through the average numbers of both hydrophobic contacts and H-bonds. Then, SMD was performed to investigate the accurate interaction energy between the monomers. By the analysis of dissociation energy, the van der Waals (vdW) force was identified as the major energy to maintain the dimer structure, which also verified the results of MD simulation. Meanwhile, 11 essential residues were discovered by the magnitude of rupture force during dissociation. Among them, residues Arg75, Glu79, Ile77, Ala88, and Trp90 were reported in a previous study using the method of mutagenesis and size exclusion chromatography, and several new important residues (Arg72, Leu74, Phe76, Gln78, Val80, and Asp91) were also identified. Interestingly, the new important residues that we discovered and the important residues that were reported are located in the opposite side of the β-sheet of the dimer.     

The influence of a protein on water dynamics in its vicinity investigated by molecular dynamics simulation

A system containing the globular protein ubiquitin and 4,197 water molecules has been used for the analysis of the influence exerted by a protein on solvent dynamics in its vicinity. Using Voronoi polyhedra, the solvent has been divided into three subsets, i.e., the first and second hydration shell, and the remaining bulk, which is hardly affected by the protein. Translational motion in the first shell is retarded by a factor of 3 in comparison to bulk. Several molecules in the first shell do not reach the diffusive regime within 100 ps. Shell-averaged orientational autocorrelation functions, which are also subject to a retardation effect, cannot be modeled by a single exponential time law, but are instead well-described by a Kohlrausch-Williams-Watts (KWW) function. The underlying distribution of single-molecule rotational correlation times is both obtained directly from the simulation and derived theoretically. The temperature dependence of reorientation is characterized by a strongly varying correlation time, but a virtually temperature-independent KWW exponent. Thus, the coupling of water structure relaxation with the respective environment, which is characteristic of each solvation shell, is hardly affected by temperature. In other words, the functional form of the distributions of single-molecule rotational correlation times is not subject to a temperature effect. On average, a correlation between reorientation and lifetimes of neighborhood relations is observed. © 1996 Wiley-Liss, Inc.     

Simulating proteins at constant pH: An approach combining molecular dynamics and Monte Carlo simulation

For the structure and function of proteins, the pH of the solution is one of the determining parameters. Current molecular dynamics (MD) simulations account for the solution pH only in a limited way by keeping each titratable site in a chosen protonation state. We present an algorithm that generates trajectories at a Boltzmann distributed ensemble of protonation states by a combination of MD and Monte Carlo (MC) simulation. The algorithm is useful for pH-dependent structural studies and to investigate in detail the titration behavior of proteins. The method is tested on the acidic residues of the protein hen egg white lysozyme. It is shown that small structural changes may have a big effect on the pK(A) values of titratable residues.     

Thermostability of protein studied by molecular dynamics simulation

The thermostability of protein thermostable cathechol 2,3-dixoygenase (TC23O) has been studied by the parallel molecular dynamics simulations. By analysis of the exponent beta, which is related to the scattering spectrum and constant-pressure heat capacity Cp, we reveal the respective contribution of a specific residue 228 proline; a specific salt bridge, Lys188N-Glu291OE1; four ions; and a different water environment to the thermostability of TC23O. The dynamic transition temperature of the mutants, Pro228Ser and Glu291Gly of the TC23O, was decreased about 10 degrees C and 19 degrees C respectively. The displacement of the four ions had no significant effect on the thermostability of TC23O. Water affects the thermostability by influencing the changes of accessible conformation to a certain extent. All these results agree with the known experimental results.     

Thermal expansion of ZnSe by molecular dynamics simulation

Thermal expansion of bulk ZnSe crystal, grown by physical vapour transport technique, is experimentally measured for various temperatures and molecular dynamics simulation using a three body Tersoff potential is also developed for the same. The resulting expansivity compares well with our experimental results and other reported values.     

Complex disruption effect of natural polyphenols on Bcl-2-Bax: molecular dynamics simulation and essential dynamics study

Apoptosis (programmed cell death) is a process by which cells died after completing physiological function or after a severe genetic damage. Apoptosis is mainly regulated by the Bcl-2 family of proteins. Anti apoptotic protein Bcl-2 prevents the Bax activation/oligomerization to form heterodimer which is responsible for release of the cytochrome c from mitochondria to the cytosol in response to death signal. Quercetin and taxifolin (natural polyphenols) efficiently bound to hydrophobic groove of Bcl-2 and altered the structure by inducing conformational changes. Taxifolin was found more efficient when compared to quercetin in terms of interaction energy and collapse of hydrophobic groove. Taxifolin and quercetin were found to dissociate the Bcl-2-Bax complex during 12?ns MD simulation. The effect of taxifolin and quercetin was, further validated by the MD simulation of ligand-unbound Bcl-2-Bax which showed stability during the simulation. Obatoclax (an inhibitor of Bcl-2) had no significant dissociation effect on Bcl-2-Bax during simulation which favored the previous experimental results and disruption effect of taxifolin and quercetin.     

Concentration effect of cimetidine with POPC bilayer: a molecular dynamics simulation study

All-atom molecular dynamics simulations have been performed on cimetidine in the presence of a palmitoyloleoylphosphatidylcholine (POPC) bilayer. The free energy profile of a single cimetidine molecule passing across POPC bilayer displays a minimum at the interface of bilayer and water. Ten cimetidine molecules were inserted into POPC bilayer to obtain an 8 mol % drug model, and molecular dynamics results showed that cimetidine molecules reside at the polar region of POPC bilayer with sulphur atoms directing to the hydrophobic region. By comparing the one drug model with 8 mol % drug model, one can see that the central barrier to cross the membrane increases while the free energy in bulk water decreases, indicating that the ability of cimetidine passing across the POPC bilayer weakens at increased concentration. In addition, the free energy minimum shifts closer to the hydrophobic core. Our results indicate that with the increased drug concentration, it is more difficult for cimetidine to enter and pass across POPC bilayer.     

Oligoglycine surface structures: molecular dynamics simulation

The full-atomic molecular dynamics (MD) simulation of adsorption mode for diantennary oligoglycines [H-Gly4-NH(CH2)5]2 onto graphite and mica surface is described. The resulting structure of adsorption layers is analyzed. The peptide second structure motives have been studied by both STRIDE (structural identification) and DSSP (dictionary of secondary structure of proteins) methods. The obtained results confirm the possibility of polyglycine II (PGII) structure formation in diantennary oligoglycine (DAOG) monolayers deposited onto graphite surface, which was earlier estimated based on atomic-force microscopy measurements.     

Exploring single-domain antibody thermostability by molecular dynamics simulation

Abstract

Single-domain antibodies also known as nanobodies are recombinant antigen-binding domains that correspond to the heavy-chain variable region of camelid antibodies. Previous experimental studies showed that the nanobodies have stable and active structures at high temperatures. In this study, the thermal stability and dynamics of nanobodies have been studied by employing molecular dynamics simulation at different temperatures. Variations in root mean square deviation, native contacts, and solvent-accessible surface area of the nanobodies during the simulation were calculated to analyze the effect of different temperatures on the overall conformation of the nanobody. Then, the thermostability mechanism of this protein was studied through calculation of dynamic cross-correlation matrix, principal component analyses, native contact analyses, and root mean square fluctuation. Our results manifest that the side chain conformation of some residues in the complementarity-determining region 3 (CDR3) and also the interaction between α-helix region of CDR3 and framework2 play a critical role to stabilize the protein at a high temperature.

Communicated by Ramaswamy H. Sarma