Structure and behavior of human α-thrombin upon ligand recognition: thermodynamic and molecular dynamics studies

Thrombin is a serine proteinase that plays a fundamental role in coagulation. In this study, we address the effects of ligand site recognition by alpha-thrombin on conformation and energetics in solution. Active site occupation induces large changes in secondary structure content in thrombin as shown by circular dichroism. Thrombin-D-Phe-Pro-Arg-chloromethyl ketone (PPACK) exhibits enhanced equilibrium and kinetic stability compared to free thrombin, whose difference is rooted in the unfolding step. Small-angle X-ray scattering (SAXS) measurements in solution reveal an overall similarity in the molecular envelope of thrombin and thrombin-PPACK, which differs from the crystal structure of thrombin. Molecular dynamics simulations performed with thrombin lead to different conformations than the one observed in the crystal structure. These data shed light on the diversity of thrombin conformers not previously observed in crystal structures with distinguished catalytic and conformational behaviors, which might have direct implications on novel strategies to design direct thrombin inhibitors.     

Structure and dynamics of two β-peptides in solution from molecular dynamics simulations validated against experiment

We have studied two different beta-peptides in methanol using explicit solvent molecular dynamics simulations and the GROMOS 53A6 force field: a heptapeptide (peptide 1) expected to form a left-handed 3(14)-helix, and a hexapeptide (peptide 2) expected to form a beta-hairpin in solution. Our analysis has focused on identifying and analyzing the stability of the dominant secondary structure conformations adopted by the peptides, as well as on comparing the experimental NOE distance upper bounds and 3J-coupling values with their counterparts calculated on the basis of the simulated ensembles. Moreover, we have critically compared the present results with the analogous results obtained with the GROMOS 45A3 (peptide 1) and 43A1 (peptide 2) force fields. We conclude that within the limits of conformational sampling employed here, the GROMOS 53A6 force field satisfactorily reproduces experimental findings regarding the behavior of short beta-peptides, with accuracy that is comparable to but not exceeding that of the previous versions of the force field. GCE legend Conformational clustering analysis of the simulated ensemble of a ss-hexapeptide with two different simulation setups (a and b). The central members of all of the clusters populating more than 5% of all of the structures are shown, together with the most dominant hydrogen bonds and the corresponding percentages of cluster members containing them.     

Structure and Conformation of the Antiviral Agent 5-Methoxymethyl-2′-deoxyuridine

Abstract

The three dimensional structure of the activiral agent, 5-methoxymethyl-2′-deoxyuridine (MmdUrd) was determined by x-ray diffraction methods. MMdUrd crystallized in space group P2₁2₁2₁ of the orthorhombic system with a = 9.166(1)A, b, = 25.348(1)Amm c = 5.270(1)A and Z = 4. The conformation of the glycosyl bond is anti (χ = 233.30), the deoxyribose ring has the C(2′)-endo envelope conformation (²E), the CH₂OH side chain has the g⁺ conformation and the methoxy group at the C(5) position is on the same side of pyrimidine plane as the 0(4′) oxygen. NMR spectroscopy was used to determine the conformation in solution. The spectra indicate that the sugar ring exists in a 60:40 equilibrium of the S- and N-states. The population of the three rotamers about the exocyclic c(4′)–C(5′) bond were estimated to be g⁺:t:g::61%:31%:8%. The correlaiton of molecular conforation with antiviral activity is discussed.     

Molecular Structure of Peptaibol Antibiotics: Solution Conformation and Crystal Structure of the Octapeptide Corresponding to the 2–9 Sequence of Emerimicins III and IV

Abstract

The infrared absorption and ¹H nuclear magnetic resonance analyses of chloroform solutions of the terminally-blocked segment corresponding to the 2–9 sequence of emerimicins III and IV, -(Aib)₃-L-Val-Gly-L-Leu-(Aib)_2-, are consistent with the presence of a 3₁₀-helical structure of high thermal stability. The crystal structure of the octapeptide, obtained by X-ray diffraction indicates the formation of a right-handed 3₁₀-helix, stabilized by six consecutive intramolecular N-H OC H-bonds, slightly distorted at the level of the L-Leu residue.     

Effects of the binding of calcium ions on the structure and dynamics of the ΦX174 virus investigated using molecular dynamics

It is known that the presence of calcium ions (Ca^2 +) is necessary for the enterobacterial virus ΦX174 to inject its DNA into the host cell, and that some mutations in the major capsid proteins lead to better survivability at higher temperatures. Our goal in the current study is to determine the physical changes in both the wild-type and mutant virus due to the binding of Ca^2 +. Thus, we performed molecular dynamics simulations of the ΦX174 major capsid protein complex with and without Ca^2 + bound. Our results show that binding of Ca^2 + leads to energetic and dynamical changes in the virus proteins. In particular, the results suggest that binding of Ca^2 + is energetically favorable and that the mutation leads to increased fluctuations of the protein complex (especially with the calcium ions bound to the complex), which may increase the rate of genome packaging and ejection for ΦX174.     

Enhancement of the chiroptical response of α-amino acids via N-substitution for molecular structure determination using vibrational circular dichroism

The chiroptical response in the form of vibrational circular dichroism (VCD) in the midinfrared region is found to be enhanced when a hydrogen of amino group of l -tryptophan is substituted with acetyl, acryloyl, or maleyl group. The order of preference for VCD enhancement is found to be acryloyl > acetyl > maleyl group. The resulting experimental VCD spectra are also found to be satisfactorily reproduced by the quantum mechanical (QM) predicted spectra. The QM predicted spectra were simulated using the conformer populations, (a) predicted by Gibbs energies and (b) optimized to maximize the similarity between experimental and predicted VCD spectra. It is found that the conformer populations predicted by Gibbs energies do not yield the maximum possible similarity between experimental and the QM predicted spectra. This work identifies the N-substitution of α-amino acids and determining the conformer populations that best reproduce the experimental spectra as two new approaches for molecular structure determination.     

Effect of Zn ion on the structure and electronic states of Aβ nonamer: molecular dynamics and ab initio molecular orbital calculations

Aggregates of amyloid-beta proteins (Aβ) have been recognised to be intimately related to pathogenesis of Alzheimer’s disease (AD). Indeed, Aβ aggregates of various sizes from dimers to fibrils were found in the brains of AD patients, and these aggregates can be self-organised. Since abnormal accumulation of metal ions such as Zn, Cu and Fe was also observed in the brains, the association between Aβ aggregations and these metal ions has been studied widely. In the present study, to elucidate the influence of Zn ions on the stability of Aβ aggregates, we performed molecular dynamics (MD) simulations and ab initio fragment molecular orbital (FMO) calculations on the Aβ nonamers with and without Zn ions and investigated the change in its structure and electronic states induced by Zn ions at atomic and electronic levels. The MD simulations revealed that Aβ nonamer cannot keep its symmetry structure, whereas Aβ nonamer with Zn ions keeps the structure. The FMO results indicated that electrostatic interactions among the charged amino-acid residues of Aβ nonamer are significantly changed by the influence of Zn ions to stabilise Aβ nonamer. These results provide useful information for proposing novel compounds, which binds specifically to Aβ and inhibits the Aβ aggregation.     

Characterization of the internal dynamics and conformational space of zinc-bound amyloid β peptides by replica-exchange molecular dynamics simulations

Amyloid β (Aβ) peptides and metal ions have been associated with the pathogenesis of Alzheimer’s disease. The conformational space of Aβ fragments of different length with and without binding of metal ions has been extensively investigated by replica-exchange molecular dynamics (REMD) simulation. However, only trajectories extracted at relatively low temperatures have been used for this analysis. The capability of REMD simulations to characterize the internal dynamics of such intrinsically disordered proteins (IDPs) as Aβ has been overlooked. In this work, we use an approach recently developed by Xue and Skrynnikov (J Am Chem Soc 133:14614–14628, 2011) to calculate NMR observables, including ¹⁵N relaxation rates and ¹⁵N–¹H nuclear Overhauser enhancement (NOE), from the high-temperature trajectory of REMD simulations for zinc-bound Aβ peptides. The time axis of the trajectory was rescaled to correct for the effect of the high temperature (408 K) compared with the experimental temperature (278 K). Near-quantitative agreement between simulated values and experimental results was obtained. When the structural properties and free-energy surfaces of zinc-bound Aβ_(1–40) and Aβ_(1–42) were compared at the physiological temperature 310 K it was found that zinc-bound Aβ_(1–42) was more rigid than Aβ_(1–40) at the C terminus, and its conformational transitions were also more preferred. The self-consistent results derived from trajectories at high and low temperatures demonstrate the capability of REMD simulations to capture the internal dynamics of IDPs.     

Structure and dynamics of α-chymotrypsin-N-trifluoroacetyl-4-fluorophenylalanine complexes

Summary Fluorine NMR lineshape, relaxation and Overhauser effect data collected at 282 and 470 MHz have been used to obtain information about the nature of complexes formed betweenN-trifluoroacetyl-4-fluorophenylalanine and the enzyme chymotrypsin. Systems involving both enantiomers have been examined as well as derivatives of these in which the aromatic ring hydrogens have been replaced by deuterium. The enzyme-induced fluorine chemical shift effects and the dynamics of molecular motions of the fluorophenyl ring at the respective binding sites appear to be similar in both complexes and, where comparable, the results are in agreement with data obtained at lower frequencies that have been reported by other workers. The dynamics of the fluoroaromatic ring in these complexes are significantly different from those observed in a closely related acylated enzyme.     

Molecular dynamics studies of simple membrane — Water interfaces: Structure and functions in the beginnings of cellular life

Molecular dynamics computer simulations of the structure and functions of a simple membrane are performed in order to examine whether membranes provide an environment capable of promoting protobiological evolution. Our model membrane is composed of glycerol 1-monooleate. It is found that the bilayer surface fluctuates in time and space, occasionally creating thinning defects in the membrane. These defects are essential for passive transport of simple ions across membranes because they reduce the Bom barrier to this process by approximately 40%. Negative ions are transferred across the bilayer more readily than positive ions due to favorable interactions with the electric field at the membrane-water interface. Passive transport of neutral molecules is, in general, more complex than predicted by the solubility-diffusion model. In particular, molecules which exhibit sufficient hydrophilicity and lipophilicity concentrate near membrane surfaces and experience interfacial resistance to transport. The membrane-water interface forms an environment suitable for heterogeneous catalysis. Several possible mechanisms leading to an increase of reaction rates at the interface are discussed. We conclude that vesicles have many properties that make them very good candidates for earliest protocells. Some potentially fruitful directions of experimental and theoretical research on this subject are proposed.     

Structure based docking and molecular dynamics studies: Peroxisome proliferator-activated receptors –α/γ dual agonists for treatment of metabolic disorders

Abstract

Diabetes is a foremost health problem globally susceptible to increased mortality and morbidity. The present therapies in the antidiabetic class have sound adverse effects and thus, emphasis on the further need to develop effective medication therapy. Peroxisome proliferator-activated receptor alpha-gamma dual approach represents an interesting target for developing novel anti-diabetic drug along with potential anti-hyperlipidimic activity. In the current study, the peroxisome proliferator-activated receptor alpha-gamma agonistic hits were screened by hierarchical virtual screening of drug like compounds followed by molecular dynamics simulation and knowledge-based structure-activity relation analysis. The key amino acid residues of binding pockets of both target proteins were acknowledged as essential and were found to be associated in the key interactions with the most potential dual hit. This dual targeted approach of structure based computational technique was undertaken to identify prevalent promising hits for both targets with binding energy and absorption distribution metabolism excretion prediction supported the analysis of their pharmacokinetic potential. In addition, stability analysis using molecular dynamics simulation of the target protein complexes was performed with the most promising dual targeted hit found in this study. Further, comparative analysis of binding site of both targets was done for the development of knowledge-based structure-activity relationship, which may useful for successful designing of dual agonistic candidates. Abbreviations ADME absorption distribution metabolism excretion

HTVS highthroughput virtual screening

MD molecular dynamics

MMGBSA molecular mechanics generalized bonn solvation accessible

PDB protein data bank

PPAR peroxisome proliferator-activated receptor

RMSD Root mean square deviation

RMSF Root mean square fluctuation

SAR structural activity relationship

SP simple precision

T2DM TypeII diabetes mellitus

XP Extra precision

Communicated by Ramaswamy H. Sarma     

Structure analysis and molecular model of the selenoenzyme glutathione peroxidase at 2.8 Å resolution

The three-dimensional structure of bovine erythrocyte glutathione peroxidase, a tetrameric enzyme containing 4 gram atoms of selenium per mole (M_r = 84,000), has been determined at 2.8 Å resolution using the multiple isomorphous replacement method. By correlation calculations in Patterson space the tetramers were shown to exhibit molecular [222] symmetry, proving the monomers to be identical or at least very similar.The monomer consists of a single polypeptide chain of 178 amino acid residues. Its shape is nearly spherical with a radius of $\text{r ≈ 19}\text{A}\text{?}$. A tentative sequence corresponding to a partially refined model (R = 0.38) is given. Each subunit is built up from a central core of two parallel and two anti-parallel strands of pleated sheet surrounded by four α-helices. One of the helices runs antiparallel to the neighbouring β-strands giving rise to a βαβ substructure, an architecture that has been found in several other proteins e.g. flavodoxin, thioredoxin, rhodanese and dehydrogenases. A comparison of the glutathione peroxidase subunit structure with thioredoxin-S₂ revealed large regions of structural resemblance. The central four-stranded β structure together with two parallel α-helices resembles nearly 80% of the thioredoxin fold.The active sites of glutathione peroxidase are located in flat depressions on the molecular surface. Probably each active centre is built up by segments from two subunits. The catalytically active selenocysteines were found at the N-terminal ends of long α-helices and are surrounded by an accumulation of aromatic side-chains. A difference Fourier map between oxidized and substrate-reduced glutathione peroxidase as well as heavy-atom binding led to the conclusion that the two-electron redox-cycle involves a reversible transition of the active-site selenium from a selenenic acid (RSeOH) to a seleninic acid (RSeOOH).     

Synthesis and characterization of MMA–NaAlg/hydroxyapatite composite and the interface analyse with molecular dynamics

A novel composite of α-methyl methacrylate (MMA) grafted sodium alginate (NaAlg) and hydroxyapatite (HA) was prepared in this study. The compositions and chemical groups of materials were investigated by infrared spectra (IR), X-ray diffraction (XRD) and nuclear magnetic resonance (NMR). The results showed that MMA has been successfully grafted with the hydroxyl group of sodium alginate. Moreover many chemical bonds existed in the composite, including the “egg-box” structure and hydrogen bonding. Meanwhile, the chemical bondings between MMA and sodium alginate partly replaced the intermolecular hydrogen bonding or the intramolecular hydrogen bonding in sodium alginate. The composite had better water contact angle than sodium alginate, indicating the strong hydrophilic character of pure sodium alginate was improved. The molecular dynamics (MD) method was used to simulate and evaluate the interaction energies based on the theoretics, which suggested that the copolymer whose every monomer grafted with one MMA had a more stable structure.     

Effect of the A30P mutation on the structural dynamics of micelle-bound αSynuclein released in water: a molecular dynamics study

Atomistic molecular dynamics simulation has been used to probe the effect of the A30P mutation on the structural dynamics of micelle-bound, helical αSynuclein when released in an aqueous environment. On the timescales simulated, the effect of the mutation on the secondary structure is restricted to local changes close to the mutation site in the N-terminal helical domain. The changes are transient, and all residues except Lys23 recover their initial structure. The local behavior due to the mutation gives rise to a global difference in the A30P mutant in the form of a permanent kink in the N-terminal helical domain.     

Ab initio and molecular dynamics studies of solid β-HMX: effects of hydrostatic pressure and high temperature

Using first-principles density functional theory and classical molecular dynamics (MD), the structural, electronic and mechanical properties of the energetic material β-HMX have been studied. The crystal structure optimised by the local density approximation calculations compares reasonably with the experimental data. Electronic band structure and density of states indicate that β-HMX is an insulator with a band gap of 3.059 eV. The pressure effect on the crystal structure and physical properties has been investigated in the range of 0–40 GPa. The crystal structure and electronic properties change slightly as the pressure increases from 0 to 2.5 GPa; when the pressure is above 2.5 GPa, further increment of the pressure results in significant changes in crystal structure. There is a larger compression along the b-axis than along the a- and c-axes. Isothermal–isobaric MD simulations on β-HMX were performed in the temperature range of 5–400 K. Phase transition at 360 K, corresponding to a volume interrupt, was found. The computed thermal expansion coefficients show anisotropic behaviour with a slightly larger expansion along the b- and c-axes than along the a-axis. In the temperature range of 5–360 K, β-HMX possesses good plasticity and its stiffness decreases with increasing the temperature.