Explicit-water Dynamics Research Articles

Adaptive Synthesis of Functional Amphiphilic Dendrons as a Novel Approach to Artificial Supramolecular Objects.

Supramolecular structures, such as micelles, liposomes, polymerosomes or dendrimerosomes, are widely studied and used as drug delivery systems. The behavior of amphiphilic building blocks strongly depends on their spatial distribution and shape of polar and nonpolar component. This report is focused on the development of new versatile synthetic protocols for amphiphilic carbosilane dendrons (amp-CS-DDNs) capable of self-assembly to regular micelles and other supramolecular objects. The presented strategy enables the fine modification of amphiphilic structure in several ways and also enables the facile connection of a desired functionality. DLS experiments demonstrated correlations between structural parameters of amp-CS-DDNs and the size of formed nanoparticles. For detailed information about the organization and spatial distribution of amp-CS-DDNs assemblies, computer simulation models were studied by using molecular dynamics in explicit water.

Explanation of the Formation of Complexes between Representatives of Oxazolidinones and HDAS-β-CD Using Molecular Modeling as a Complementary Technique to cEKC and NMR.

Molecular modeling (MM) results for tedizolid and radezolid with heptakis-(2,3-diacetyl-6-sulfo)-β-cyclodextrin (HDAS-β-CD) are presented and compared with the results previously obtained for linezolid and sutezolid. The mechanism of interaction of chiral oxazolidinone ligands belonging to a new class of antibacterial agents, such as linezolid, tedizolid, radezolid, and sutezolid, with HDAS-β-CD based on capillary electrokinetic chromatography (cEKC), nuclear magnetic resonance (NMR) spectroscopy, and MM methods was described. Principles of chiral separation of oxazolidinone analogues using charged single isomer derivatives of cyclodextrin by the cEKC method were presented, including the selection of the optimal chiral selector and separation conditions, complex stoichiometry, and binding constants, which provided a comprehensive basis for MM studies. In turn, NMR provided, where possible, direct information on the geometry of the inclusion complexes and also provided the necessary structural information to validate the MM calculations. Consequently, MM contributed to the understanding of the structure of diastereomeric complexes, the thermodynamics of complexation, and the visualization of their structures. The most probable mean geometries of the studied supramolecular complexes and their dynamics (geometry changes over time) were determined by molecular dynamics methods. Oxazolidinone ligands have been shown to complex mainly the inner part of cyclodextrin, while the external binding is less privileged, which is consistent with the conclusions of the NMR studies. Enthalpy values of binding of complexes were calculated using long-term molecular dynamics in explicit water as well as using molecular mechanics, the Poisson–Boltzmann or generalized Born, and surface area continuum solvation (MM/PBSA and MM/GBSA) methods. Computational methods predicted the effect of changes in pH and composition of the solution on the strength and complexation process, and it adapted the conditions selected as optimal during the cEKC study. By changing the dielectric constant in the MM/PBSA and MM/GBSA calculations, the effect of changing the solution to methanol/acetonitrile was investigated. A fairly successful attempt was made to predict the chiral separation of the oxazolidinones using the modified cyclodextrin by computational methods.

CuFeO2–Water Interface under Illumination: Structural, Electronic, and Catalytic Implications for the Hydrogen Evolution Reaction

CuFeO2 is a p-type semiconductor that has been recently identified as a promising photocathode material for photoelectrochemical water splitting. CuFeO2 can absorb solar light and promote the hydrogen evolution reaction (HER), even though the photocurrents achieved so far are still well below the theoretical upper limit. While several experimental and theoretical works have provided a detailed characterization of the bulk properties of this material, surfaces have been largely unexplored. In this work, we perform first-principles simulations based on DFT to investigate the structure, electronic properties, and thermodynamic stability of CuFeO2 surfaces both in vacuum and in an electrochemical environment. To estimate the alignment of the band edges on the electrochemical scale, we perform ab initio molecular dynamics in explicit water, unraveling the structure of the solid/liquid interface for various surface terminations. We consider the system both in the dark and under illumination, showing that light absorption can induce partial reduction of the surface, giving rise to states in the gap that can pin the Fermi level, in agreement with recent measurements. Using the free energy of adsorption of atomic hydrogen as a descriptor of the catalytic activity for the HER, we show that hydride species formed at oxygen vacancies can be highly active and could therefore be an intermediate of reaction.

The Hypervariable Loops of Free TCRs Sample Multiple Distinct Metastable Conformations in Solution.

CD4+ and CD8+ αβ T cell antigen recognition is determined by the interaction between the TCR Complementarity Determining Region (CDR) loops and the peptide-presenting MHC complex. These T cells are known for their ability to recognize multiple pMHC complexes, and for a necessary promiscuity that is required for their selection and function in the periphery. Crystallographic studies have previously elucidated the role of structural interactions in TCR engagement, but our understanding of the dynamic process that occurs during TCR binding is limited. To better understand the dynamic states that exist for TCR CDR loops in solution, and how this relates to their states when in complex with pMHC, we simulated the 2C T cell receptor in solution using all-atom molecular dynamics in explicit water and constructed a Markov State Model for each of the CDR3α and CDR3β loops. These models reveal multiple metastable states for the CDR3 loops in solution. Simulation data and metastable states reproduce known CDR3β crystal conformations, and reveal several novel conformations suggesting that CDR3β bound states are the result of search processes from nearby pre-existing equilibrium conformational states. Similar simulations of the invariant, Type I Natural Killer T cell receptor NKT15, which engages the monomorphic, MHC-like CD1d ligand, demonstrate that iNKT TCRs also have distinct states, but comparatively restricted degrees of motion.

An atomic charge model for graphene oxide for exploring its bioadhesive properties in explicit water.

Graphene Oxide (GO) has been shown to exhibit properties that are useful in applications such as biomedical imaging, biological sensors, and drug delivery. The binding properties of biomolecules at the surface of GO can provide insight into the potential biocompatibility of GO. Here we assess the intrinsic affinity of amino acids to GO by simulating their adsorption onto a GO surface. The simulation is done using Amber03 force-field molecular dynamics in explicit water. The emphasis is placed on developing an atomic charge model for GO. The adsorption energies are computed using atomic charges obtained from an ab initio electrostatic potential based method. The charges reported here are suitable for simulating peptide adsorption to GO.

Ensemble fits of restrained peptides’ conformational equilibria to NMR data. Dependence on force fields: AMBER/8 ff03 versus ECEPP/3

Two variants of NMR-based conformational analyses of flexible peptides are compared using two examples meeting the formula Tyr- d-Daa-Phe-Daa-NH 2 (Daa = diamino acid): 1 combining d-Dab 2 (α,γ-diaminobutyryl) with Lys 4, and 2 – d-Dap 2 (α,β-diaminopropionyl) with Orn 4. The ω-amino groups of d-Daa 2 and Daa 4 are coupled with 〉C O into the urea, restraining 1 and 2 with 16- and 14-membered rings and leading to potent and impotent μ/δ opioid peptides, respectively. To the current task, we took from an earlier work ( Filip et al, J. Pept. Sci. 11 (2005) 347–352) the NMR NOE- and J-data in H 2O/D 2O; and the selection of the ensembles of 1 and 2, 822 and 788 conformational families, respectively, obtained by using the EDMC/ECEPP3 method. Here, we generated ensembles of 1 and 2 using AMBER molecular dynamics in explicit water to eventually selected 686 and 761 conformers for 1 and 2, respectively. We did numbers of fits for both types of the conformational ensembles of 1 and 2 to their NOE- and J-data using a common method i.e. maximum entropy approach ( Groth et al, J. Biomol. NMR 15 (1999) 315–330). Both types of the well structurally diversified ensembles fit to quite different equilibria in regressions to common experimental NOE- and J-restraints using maximum entropy principle, which is a disappointing message. Intriguing is startlingly small standard deviation in J-couplings: σ J NH α H ≈ 0.01 Hz for LES-MD/AMBER ensemble, contrary to σ J NH α H = 0.8 − 1.1 Hz for the EDMC/ECEPP ensemble, over the wide range of entropy, i.e. relatively insensitive to it. A similar feature is not the case when comparing σ NOE in both methods. Hence, at minute entropy contributions, it follows that J does or does not transpose “overfitted” into the final σ J in the AMBER or ECEPP ensemble, respectively. Could this be an effect of softness of the AMBER flexible-valence force field compared to ECEPP rigid-geometry, and its effect on ensemble sampling? We do not know an answer.

Solution NMR structure of the plasmid‐encoded fimbriae regulatory protein PefI from Salmonella enterica serovar Typhimurium

The surfaces of many bacteria feature pili or fimbriae, proteinaceous filaments that play an integral role in the adhesion of bacteria to host cells and, hence, in their pathogenicity.1,2 These extracellular structures are key virulence factors and potential targets for antibacterial drug and vaccine development. In certain Salmonella enterobacteria strains, one such class of fimbriae, the virulence plasmid-encoded fimbriae encoded by the pef operon,3,4 has been shown to be important for adhesion to murine small intestine and fluid accumulation,5 as well as biofilm formation on eukaryotic cell surfaces.6 Transcription of the pef operon, induced under acidic conditions, is modulated by the PefI regulatory protein.7 PefI represses plasmid-encoded fimbrial protein production by activating the leucine-responsive regulatory protein (Lrp) mediated inhibition of DNA methylation within the pef promoter region.7 PefI expression also inhibits transcription of flagellar proteins and consequently cell motility.8 Here we present the solution NMR structure of the 70-residue PefI transcription regulator from Salmonella enterica serovar Typhimurium LT2 [UniProtKB/TrEMBL ID, Q04822_SALTY; NESG ID, StR82; hereafter referred to as stPefI], a member of the FaeA-like protein domain family (Pfam identifier, PF04703). The sequence alignment of stPefI with its functional homolog in Escherichia coli, PapI, is shown in Fig. 1(A). We demonstrate that the structure of stPefI adopts a winged helix-turn-helix motif,9,10 consistent with its role as a DNA-binding transcriptional regulator. Moreover, in spite of their relatively low sequence identity (29%), the structure of stPefI is highly similar to that of E. coli PapI,11 which activates the expression of pyelonephritis-associated pili.12,13 Fig. 1 (A) Structure-based sequence alignment of stPefI and E. coli PapI (ecPapI). The sequence numbering for stPefI and the secondary structural elements found in its solution NMR structure (PDB ID, 2JT1) are shown above the alignment; t, turn and l, loop. ... Materials and Methods Isotopically enriched samples of stPefI were cloned, expressed, and purified, and the sample buffer was optimized for NMR studies following standard protocols of the NESG consortium;14,15 see Supplementary Material for a complete description of the methods used in this work. Briefly, samples of [U-13C,15N]- and [U-5%-13C,100%-15N]-stPefI for NMR spectroscopy were concentrated by ultracentrifugation to 0.5 to 0.7 mM in 95% H2O / 5% 2H2O solution containing 20 mM ammonium acetate, 450 mM NaCl, 10 mM DTT, 5 mM CaCl2 at pH 5.5. Analytical gel filtration chromatography, static light scattering (Supplementary Fig. S1) and one-dimensional 15N T1 and T2 relaxation data (Supplementary Fig. S2) demonstrate that the protein is monomeric under the conditions used in the NMR studies. All NMR data for resonance assignment and structure determination were collected at 20 °C on Bruker AVANCE 600 and 800 MHz spectrometers equipped with conventional 5-mm TXI probes, and a Varian INOVA 600 MHz instrument with a 5-mm HCN cold probe, and referenced to internal DSS (2,2-dimethyl-2-silapentane-5-sulfonic acid). Complete 1H, 13C, and 15N resonance assignments for stPefI were determined using conventional triple resonance NMR methods, with automated backbone assignment made by AutoAssign 2.4.0,16 followed by manual side chain assignment. Resonance assignments were validated using the Assignment Validation Suite (AVS) software package17 and deposited in the BioMagResDB (BMRB accession number, 15386). The solution NMR structure of stPefI was calculated using CYANA 2.1,18,19 and the 20 structures with lowest target function out of 100 in the final cycle were further refined by restrained molecular dynamics in explicit water using CNS 1.2.20,21 Structural statistics and global structure quality factors were computed using the PSVS 1.4 suite of structure quality assessment programs.22 The global goodness-of-fit of the final structure ensemble with the NOESY peak list data was determined using the RPF analysis program.23 The final ensemble of 20 models (excluding the C-terminal His6) were deposited into the Protein Data Bank (PDB ID, 2JT1). All structure figures were rendered using PyMOL (www.pymol.org).

Thermodynamic Perspective on the Dock−Lock Growth Mechanism of Amyloid Fibrils

The mechanism of addition of a soluble unstructured monomer to a preformed ordered amyloid fibril is a complex process. On the basis of the kinetics of monomer disassociation of Abeta(1-40) from the amyloid fibril, it has been suggested that deposition is a multistep process involving a rapid reversible association of the unstructured monomer to the fibril surface (docking) followed by a slower conformational rearrangement leading to the incorporation onto the underlying fibril lattice (locking). By exploiting the vast time scale separation between the dock and lock processes and using molecular dynamics simulation of deposition of the disordered peptide fragment (35)MVGGVV(40) from the Abeta peptide onto the fibril with known crystal structure, we provide a thermodynamic basis for the dock-lock mechanism of fibril growth. Free energy profiles, computed using implicit solvent model and enhanced sampling methods with the distance (delta(C)) between the center of mass of the peptide and the fibril surface as the order parameter, show three distinct basins of attraction. When delta(C) is large, the monomer is compact and unstructured and the favorable interactions with the fibril results in stretching of the peptide at delta(C) approximately 13 A. As delta(C) is further decreased, the peptide docks onto the fibril surface with a structure that is determined by a balance between intrapeptide and peptide fibril interactions. At delta(C) approximately 4 A, a value that is commensurate with the spacing between beta-strands in the fibril, the monomer expands and locks onto the fibril. Using simulations with implicit solvent model and all atom molecular dynamics in explicit water, we show that the locked monomer, which interacts with the underlying fibril, undergoes substantial conformational fluctuations and is not stable. The cosolutes urea and TMAO destabilize the unbound phase and stabilize the docked phase. Interestingly, small crowding particles enhance the stability of the fibril-bound monomer only marginally. We predict that the experimentally measurable critical monomer concentration, C(R), at which the soluble unbound monomer is in equilibrium with the ordered fibril, increases sharply as temperature is increased under all solution conditions.

Molecular dynamics simulation of antimicrobial peptide arenicin‐2: β‐Hairpin stabilization by noncovalent interactions

Arenicin-2 is a 21 residue antimicrobial cyclic peptide, possessing one disulphide bond between residues Cys(3) and Cys(20). NMR and CD studies suggested that the structure of arenicin-2 in water represented a well formed, but highly twisted beta-hairpin. To investigate the spatial arrangement of the peptide side chains and to get a clear view of its possible amphipathic properties we performed molecular dynamics in explicit water. Four independent trajectories, 50 ns in length, were produced, starting from various initial conformations or by applying different simulation conditions. Arenicin-2 retained its beta-hairpin structure during simulations, although the residues close to strand ends were found to escape from the ideal hairpin conformation. The type I' beta-turn connecting the two strands fluctuated between type IV and II' beta-turn. Conversely, the right-handed twist of the beta-hairpin was well conserved with average twist value 203 degrees +/- 19 degrees per eight residues. Several nonbonded interactions, like hydrophobic interactions between aliphatic side chains, cation/pi-aromatic interactions, CH...pi aromatic bond and water bridges, contributed to the hairpin stabilization.

Structural Basis of the Preferential Binding for Globo-Series Glycosphingolipids Displayed by Pseudomonas aeruginosa Lectin I

The opportunistic pathogen Pseudomonas aeruginosa contains several carbohydrate-binding proteins, among which is the P. aeruginosa lectin I (PA-IL), which displays affinity for α-galactosylated glycans. Glycan arrays were screened and demonstrated stronger binding of PA-IL toward αGal1–4βGal-terminating structures and weaker binding to αGal1–3βGal ones in order to determine which human glycoconjugates could play a role in the carbohydrate-mediated adhesion of the bacteria. This was confirmed in vivo by testing the binding of the lectin to Burkitt lymphoma cells that present large amounts of globotriaosylceramide antigen Gb3/CD77/P k. Trisaccharide moieties of Gb3 (αGal1–4βGal1–4Glc) and isoglobotriaosylceramide (αGal1–3βGal1–4Glc) were tested by titration microcalorimetry, and both displayed similar affinity to PA-IL in solution. The crystal structure of PA-IL complexed to αGal1–3βGal1–4Glc trisaccharide has been solved at 1.9-Ǻ resolution and revealed how the second galactose residue makes specific contacts with the protein surface. Molecular modeling studies were performed in order to compare the binding mode of PA-IL toward αGal1–3Gal with that toward αGal1–4Gal. Docking studies demonstrated that αGal1–4Gal creates another network of contacts for achieving a very similar affinity, and 10-ns molecular dynamics in explicit water allowed for analyzing the flexibility of each disaccharide ligand in the protein binding site. The higher affinity observed for binding to Gb3 epitope, both in vivo and on glycan array, is likely related to the presentation effect of the oligosaccharide on a surface, since only the Gb3 glycosphingolipid geometry is fully compatible with parallel insertion of neighboring trisaccharide heads in two binding sites of the same tetramer of PA-IL.

Computational Study of the Mechanism and the Relative Free Energies of Binding of Anticholesteremic Inhibitors to Squalene-Hopene Cyclase

The prokaryotic monotopic membrane protein squalene-hopene cyclase (SHC) is homologous to a human enzyme responsible for cholesterol formation. Using molecular dynamics in explicit water, a single monomer of SHC was simulated using the GROMOS 45A3 force field, once in complex with an inhibitor and once in an uncomplexed form. The protein exhibits significant stability on the level of secondary and tertiary structure even outside of its native membrane environment. Analysis of the fluctuations of the complexed and the uncomplexed SHC confirms the previously made suggestions for the ligand entrance channel and reveals some of its novel dynamical features that might be of functional importance. To examine the potential of computationally designing SHC ligands and study their thermodynamics of binding, the relative free energies of binding of a series of structurally similar anticholesteremic inhibitors of SHC were calculated using single-step perturbation (SSP) and thermodynamic integration (TI) techniques. While neither technique succeeds in quantitatively matching the relatively small experimental values, TI qualitatively reproduces the relative order of the experimental affinities, but SSP does not. Detailed comparisons and potential reasons for this are given.

Polarizable atomic multipole solutes in a Poisson-Boltzmann continuum.

Modeling the change in the electrostatics of organic molecules upon moving from vacuum into solvent, due to polarization, has long been an interesting problem. In vacuum, experimental values for the dipole moments and polarizabilities of small, rigid molecules are known to high accuracy; however, it has generally been difficult to determine these quantities for a polar molecule in water. A theoretical approach introduced by Onsager [J. Am. Chem. Soc. 58, 1486 (1936)] used vacuum properties of small molecules, including polarizability, dipole moment, and size, to predict experimentally known permittivities of neat liquids via the Poisson equation. Since this important advance in understanding the condensed phase, a large number of computational methods have been developed to study solutes embedded in a continuum via numerical solutions to the Poisson-Boltzmann equation. Only recently have the classical force fields used for studying biomolecules begun to include explicit polarization in their functional forms. Here the authors describe the theory underlying a newly developed polarizable multipole Poisson-Boltzmann (PMPB) continuum electrostatics model, which builds on the atomic multipole optimized energetics for biomolecular applications (AMOEBA) force field. As an application of the PMPB methodology, results are presented for several small folded proteins studied by molecular dynamics in explicit water as well as embedded in the PMPB continuum. The dipole moment of each protein increased on average by a factor of 1.27 in explicit AMOEBA water and 1.26 in continuum solvent. The essentially identical electrostatic response in both models suggests that PMPB electrostatics offers an efficient alternative to sampling explicit solvent molecules for a variety of interesting applications, including binding energies, conformational analysis, and pK(a) prediction. Introduction of 150 mM salt lowered the electrostatic solvation energy between 2 and 13 kcalmole, depending on the formal charge of the protein, but had only a small influence on dipole moments.